Nicotinoyl riboside compositions and methods of use

ABSTRACT

The invention relates to compositions of nicotinoyl ribosides and nicotinamide riboside derivatives and their methods of use. In some embodiments, the invention relates to methods of making nicotinoyl ribosides. In some embodiments, the invention relates to pharmaceutical compositions and nutritional supplements containing a nicotinoyl riboside. In further embodiments, the invention relates to methods of using nicotinoyl ribosides and nicotinamide riboside derivatives that promote the increase of intracellular levels of nicotinamide adenine dinucleotide (NAD+) in cells and tissues for improving cell and tissue survival.

CROSS-REFERENCE TO RELATED APPLICATIONS

This patent application is a continuation of copending U.S. patentapplication Ser. No. 16/888,336, filed May 29, 2020, which is acontinuation of Ser. No. 16/277,741, filed Feb. 15, 2019, now issued asU.S. Pat. No. 10,668,096, which is a continuation of U.S. patentapplication Ser. No. 15/137,818, filed Apr. 25, 2016, now issued as U.S.Pat. No. 10,206,940, which is a continuation of U.S. patent applicationSer. No. 14/543,439, filed Nov. 17, 2014, now issued as U.S. Pat. No.9,321,797, which is a continuation of U.S. patent application Ser. No.13/351,411, filed Jan. 17, 2012, now issued as U.S. Pat. No. 9,000,147,which is a divisional of U.S. patent application Ser. No. 11/601,714,filed Nov. 17, 2006, now issued as U.S. Pat. No. 8,106,184, which claimsthe benefit of U.S. Provisional Application No. 60/738,081, filed Nov.18, 2005, the disclosures of which are hereby incorporated by referencein their entireties and for all purposes.

FIELD OF INVENTION

The invention relates to compositions of nicotinoyl ribosides andnicotinamide riboside derivatives and their methods of use. In someembodiments, the invention relates to methods of making nicotinoylribosides. In some embodiments, the invention relates to pharmaceuticalcompositions and nutritional supplements containing a nicotinoylriboside. In further embodiments, the invention relates to methods ofusing nicotinoyl ribosides and nicotinamide riboside derivatives thatpromote the increase of intracellular levels of nicotinamide adeninedinucleotide (NAD+) in cells and tissues for improving cell and tissuesurvival.

BACKGROUND

Nicotinamide adenine dinucleotide (NAD+) is a natural coenzyme thatfunctions as an intermediary in cellular oxidation and reductionreactions as well as an ADP-ribosyltransferase substrate. Alteringintracellular NAD+ levels can improve the health of a cell, butintroduction of compounds that enter NAD+ metabolic pathways can alsoprove toxic to cells. For example, benzamide riboside (BAR) is awell-known antitumor agent. BR is a prodrug that can be phosphorylatedto its 5′-monophosphate and then converted to its active metabolitebenzamide adenine dinucleotide (BAD). That metabolite is an activeanalogue of NAD+ and an inhibitor of inosine 5′-monophosphatedehydrogenase (IMPHD). IMPDH is linked to malignant transformations. BARshows selective sensitivity to central nervous system and leukemic celllines. However, BAD also inhibits other dehydrogenases, such as malatedehydrogenase and glutamic acid dehydrogenase which may cause adverseeffects if used as a therapeutic. Thus, there is a need to identifycompositions that are capable of improving the health of damaged ordiseased cells preferably by altering intracellular NAD+ levels and thatdo not have adverse effects if the compositions are giventherapeutically.

SUMMARY OF INVENTION

The invention relates to compositions of nicotinoyl ribosides andnicotinamide riboside derivatives and their methods of use. In someembodiments, the invention relates to methods of making nicotinoylribosides including nicotinamide riboside. In some embodiments, theinvention relates to pharmaceutical compositions and nutritionalsupplements containing a nicotinoyl riboside. In further embodiments,the invention relates to methods of using nicotinoyl ribosides andnicotinamide riboside derivatives that promote the increase ofintracellular levels nicotinamide adenine dinucleotide (NAD+) in cellsand tissues for improving cell and tissue survival.

In some embodiments the invention relates to a compound having theformula:

and salt complexes thereof wherein R is selected from the groupconsisting of alkylamino, substituted alkylamino, dialkylamino,substituted dialkylamino, alkyloxy, substituted alkyloxy, aryloxy,substituted aryloxy, hydroxylamino, substituted hydroxylamino,O-alkyloxyamino, substituted O-alkyloxyamino, aminooxy, substitutedaminoxy, N-alkylaminooxy, substituted N-alkylaminoxy, hydrazino,substituted hydrazino, alkylhydrazino, and substituted alkylhydrazino.

In additional embodiments, the invention relates to a compound havingthe following formula:

and salt complexes thereof wherein, R¹ is alkyloxy, substitutedalkyloxy, aryloxy, substituted aryloxy, arylthio, substituted arylthio,alkylthio, substituted alkylthio, aminooxy, substituted aminoxy,N-alkylaminooxy, and substituted N-alkylaminooxy; and R², R³, and R⁴ arethe same or different and, at each occurrence, independently acyl orsubstituted acyl.

In further embodiments, the invention relates to a compound having thefollowing formula:

and salt complexes thereof wherein, R¹ is alkylamino, substitutedalkylamino, dialkylamino, substituted dialkylamino, hydrazine,substituted hydrazino, N-alkylaminohydrazino, hydroxylamino, substitutedhydroxylamino, O-alkyloxyamino, or substituted O-alkyloxyamino; R², R³,and R⁴ are the same or different and, at each occurrence, independentlyacyl or substituted acyl.

In additional embodiments, the invention relates to a method of making acompound having Formula I:

wherein, R¹ is amino, alkylamin, substituted alkylamino, dialkylamino,substituted dialkylamino, alkyloxy, substituted alkyloxy, aryloxy,substituted aryloxy, hydroxylamino, substituted hydroxylamino,O-alkyloxyamino, substituted O-alkyloxyamino, aminooxy, substitutedaminoxy, N-alkylaminooxy, substituted N-alkylaminoxy, hydrazino,substituted hydrazino, alkylhydrazino, and substituted alkylhydrazinocomprising: i) mixing a) tetra-acyl or substituted acyl ribofuranose andb) a compound having formula II:

wherein, R² is alkyloxy, substituted alkyloxy, aryloxy, substitutedaryloxy, alkylthiol, substituted alkylthiol, arylthiol, substitutedarylthiol, aminooxy, substituted aminooxy, N-alkylaminooxy orsubstituted N-alkylaminooxy; under conditions such that a compound offormula III:

wherein, R² is alkyloxy, substituted alkyloxy, aryloxy, substitutedaryloxy, alkylthiol, substituted alkylthiol, arylthiol, substitutedarylthiol, aminooxy, substituted aminooxy, N-alkylaminooxy orsubstituted N-alkyl aminooxy, and R³, R⁴ and R⁵ are the same ordifferent and, at each occurrence, independently acyl or substitutedacyl is formed; and ii) mixing the compound of formula III with ammonia,hydrazine, substituted hydrazine, alkylhydrazine, substitutedalkylhydrazine, hydroxylamine, substituted hydroxylamine,O-alkyloxyamine, substituted O-alkyloxylamine, N-alkylaminohydroxide,substituted N-alkylaminohydroxide, alkylamine, substituted alkylamine,dialkylamine, substituted dialkylamine, alkoxide, or substitutedalkoxide under conditions such that a compound of formula I is formed.In further embodiments, the conditions of making compound I fromcompound III are in a halogenated alkyl alcohol solvent at a temperaturebelow room temperature. In further embodiments, said solvent istrifluoromethanol.

In some embodiments, the invention relates to a compound having thefollowing formula:

and salt complexes thereof wherein, R¹ is alkyloxy, substitutedalkyloxy, aryloxy, substituted aryloxy, arylthio, substituted arylthio,alkylthio, or substituted alkylthio; and R², R³, and R⁴ are the same ordifferent and, at each occurrence, independently acyl or substitutedacyl.

In some embodiments, the invention relates to a method of making acompound having Formula I:

wherein, R¹ is amine, substituted amine, alkylamine, substitutedalkylamine, dialkylamine, substituted dialkylamine, alkyloxy, orsubstituted alkyloxy, comprising: i) mixing a) tetra-acyl or substitutedacyl ribofuranose, preferably tetra-acetyl ribofuranose, and b) acompound having formula II:

wherein, R² is alkyloxy, substituted alkyloxy, aryloxy, substitutedaryloxy, alkylthiol, substituted alkylthiol, arylthiol or substitutedarylthiol under conditions such that a compound of formula III:

wherein, R² is alkyloxy, substituted alkyloxy, aryloxy, substitutedaryloxy, alkylthiol, substituted alkylthiol, arylthiol or substitutedarylthiol, and R³, R⁴ and R⁵ are the same or different and, at eachoccurrence, independently acyl or substituted acyl is formed; and ii)mixing the compound of formula III with ammonia, substituted ammoniaalkylamine, substituted alkylamine, dialkylamine, substituteddialkylamine, alkoxide, or substituted alkoxide under conditions suchthat a compound of formula I is formed.

In some embodiments, the invention relates to a compound having theformula:

and salt complexes thereof wherein R is selected from the groupconsisting of alkylamine, substituted alkylamine, dialkylamine,substituted dialkylamine, alkyloxy, and substituted alkyloxy.

In additional embodiments, the invention relates to a compound havingthe following formula:

and salt complexes thereof wherein, R¹ is alkyloxy or substitutedalkyloxy, or aryloxy or alkylthio or arylthio; and R², R³, and R⁴ arethe same or different and, at each occurrence, independently acyl orsubstituted acyl.

In some embodiments, the invention relates to a compound having thefollowing formula:

and salt complexes thereof wherein, R¹ is aminoalkyl, substituted aminoalkyl, diaminoalkyl, or substituted diaminoalkyl and R², R³, and R⁴ arethe same or different and, at each occurrence, independently acyl orsubstituted acyl.

In some embodiments, the invention relates to a method of making acompound having Formula I:

wherein, R is NH₂, alkylamino, substituted alkylamino, dialkylamino,substituted dialkylamino, alkyloxy, and substituted alkyloxy,comprising: i) mixing tetra-acyl ribofuranose, preferably tetra-acetylribofuranose, and alkyl or aryl pyridine-3-carboxylate ester orthioester compound under conditions such that a compound of formula II:

is formed where R² is —O-alkyl, —O-aryl, —S-alkyl or —S-aryl and, R³, R⁴and R⁵ are the same or different and, at each occurrence, independentlyacyl and iii) mixing the compound of formula II with an alkylamine,substituted alkylamine, dialkylamine, substituted dialkylamine,alkoxide, and substituted alkoxide under conditions such that a compoundof formula I is formed.

In some embodiments, the invention relates to making alkyl1-[3,4-diacetyloxy-5-(acetyloxymethyl)oxolan-2-yl]-pyridine-3-carboxylate,preferably alkyl1-[3,4-diacetyloxy-5-(acetyloxymethyl)oxolan-2-yl]-pyridine-3-carboxylate,comprising mixing a composition consisting essentially of1,2,3,5-tetra-O-acetyl-beta-D-ribofuranose, alkylpyridine-3-carboxylate, and trimethylsilyl trifluoromethanesulfonateunder conditions such that alkyl1-[3,4-diacetyloxy-5-(acetyloxymethyl)oxolan-2-yl]-pyridine-3-carboxylateis formed.

In some embodiments, the invention relates to a method of making N-alkylor N,N dialkyl1-[3,4-dihydroxy-5-(hydroxymethyl)oxolan-2-yl]pyridine-5-carboxamidecomprising mixing alkyl1-[3,4-diacetyloxy-5-(acetyloxymethyl)oxolan-2-yl]-pyridine-3-carboxylatewith an primary or secondary amine under conditions such that N-alkyl orN,N dialkyl1-[3,4-dihydroxy-5-(hydroxymethyl)oxolan-2-yl]pyridine-5-carboxamide isformed.

In some embodiments, the invention relates to a method of makingO-Alkyl²1-[3,4-dihydroxy-5-(hydroxymethyl)oxolan-2-yl]pyridine-5-carboxylatecomprising mixingO-Alkyl¹-[3,4-dihydroxy-5-(hydroxymethyl)oxolan-2-yl]pyridine-5-carboxylateand sodium alkyl² oxide under conditions such that O-Alky²1-[3,4-dihydroxy-5-(hydroxymethyl)oxolan-2-yl]pyridine-5-carboxylate isformed.

In some embodiments, the invention relates to a substituted orunsubstituted compound capable of inhibiting inosine 5′-monophosphatedehydrogenase having the formula:

wherein, R is alkylamino, substituted alkylamino, dialkylamino,substituted dialkylamino, alkyloxy, and substituted alkyloxy. In furtherembodiments, R is —OMe, —OEt, —OCH₂CH₂OH, —NHMe, —NHEt, —NHCH₂CH₂OH,—NHCH₂CH═CH₂, —NHCH(CH₃)₂, —NHCH₂CH₂NH₂, —NHcyclopropyl, —N(CH₃)₂, or Nsubstituted pyrrolidinyl. In further embodiments, the compound isobtained in a purified form.

In some embodiments, the invention relates to a compound having thefollowing formula.

wherein, R¹ is alkylamino, substituted alkylamino, dialkylamino, orsubstituted dialkylamino; n is 1 or 2; and R², R³, and R⁴ are are thesame or different and, at each occurrence, independently acyl orsubstituted acyl. In additional embodiments, R¹ is not —NHMe, —NHEt, or—NEt₂.

In some embodiments, the invention relates to a purified compound havingthe following formula:

wherein, R¹ is alkyloxy or substituted alkyloxy, preferably methoxy andethoxy and R², n is one or two; and R³, and R⁴ are the same or differentand, at each occurrence, independently acyl or substituted acyl,preferably acetyl and benzoyl. In additional embodiments, R¹ is not —OMeor —OEt.

In some embodiments, the invention relates to a method of making annicotinamide derivative comprising: i) mixing tetra-acyl ribofuranose,preferably tetra-acetyl ribofuranose and alkyl or arylpyridine-3-carboxylate ester or thioester compound under conditions suchthat a second compound having the following formula:

is formed wherein R² is —O-alkyl, —O-aryl, —S-alkyl or —S-aryl and, R³,R⁴ and R⁵ are the same or different and, at each occurrence,independently acyl and iii) mixing said second compound with H—R,wherein R is —OMe, —OEt, —OCH₂CH₂OH, —NHMe, —NHEt, —NHCH₂CH₂OH,—NHCH₂CH═CH₂, —NHCH(CH₃)₂, —NHCH₂CH₂NH₂, —NHcyclopropyl, —N(CH₃)₂, or Nsubstituted pyrrolidinyl; under conditions such that a compound havingthe following formula:

is formed.

In some embodiments, the invention relates to compounds disclosed hereinand salt complexes thereof. In further embodiment, said salt complexesare pharmaceutical preparations or nutritional supplement.

In other embodiments, the composition comprises a nicotinoyl ribosidecomponent comprising greater than 80%, or greater than 90%, 95% or 99%of a nicotinoyl riboside compound disclosed herein.

In some embodiments, the invention relates to a method of managing orimproving the health of cells, preferably stem cells, more preferablyembryonic stem cells comprising: i) providing a) a compositioncomprising nicotinoyl riboside substituted through the carbonyl to forma substituted or unsubstituted ester or alkyl or dialkyl amide disclosedherein and b) a cell; and ii) mixing said composition and said cellunder conditions such that the heath of said cell is managed orimproved.

In some embodiments, the invention relates to a growth medium containinga nicotinoyl riboside or a nicotinamide derivative disclosed herein. Infurther embodiments, the invention relates to methods of usingnicotinoyl ribosides and nicotinamide riboside derivatives forincreasing NAD+ levels in cells and tissues and for improving cell andtissue survival. In further embodiments said cells are embryonic stemcells and said tissue is a blastocyst comprising embryonic stem cells.

In some embodiment, the invention relates to a method of managing,preventing or treating cancer preferably leukemia comprising i)providing a) a subject having symptoms of cancer preferably leukemia andb) a nicotinoyl riboside derivative disclosed herein and ii)administering said derivative to said subject. In further embodiments, anicotinoyl riboside or nicotinamide derivatives that inhibits inosine5′-monophosphate dehydrogenase.

In some embodiments, the invention relates to a method of managing orimproving the health of cells, preferably stem cells, more preferablyembryonic stem cells comprising: i) providing a) a compositioncomprising compound having the following formula:

wherein R is —OMe, —OEt, —OCH₂CH₂OH, —NHMe, —NHEt, —NHCH₂CH₂OH,—NHCH₂CH═CH₂, —NHCH(CH₃)₂, —NHCH₂CH₂NH₂, —NHcyclopropyl, —N(CH₃)₂, or Nsubstituted pyrrolidinyl and b) a cell; and ii) mixing said compositionand said cell under conditions such that the heath of said cell ismanaged or improved.

In additional embodiments, the invention relates to a growth mediumcontaining a compound having the following formula:

wherein R is —OMe, —OEt, —OCH₂CH₂OH, —NHMe, —NHEt, —NHCH₂CH₂OH,—NHCH₂CH═CH₂, —NHCH(CH₃)₂, —NHCH₂CH₂NH₂, —NHcyclopropyl, —N(CH₃)₂, or Nsubstituted pyrrolidinyl.

In some embodiment, the invention relates to a method of managing,preventing or treating cancer preferably leukemia comprising i)providing a) a subject having symptoms of cancer preferably leukemia andb) containing a compound having the following formula:

wherein R is —OMe, —OEt, —OCH₂CH₂OH, —NHMe, —NHEt, —NHCH₂CH₂OH,—NHCH₂CH═CH₂, —NHCH(CH₃)₂, —NHCH₂CH₂NH₂, —NHcyclopropyl, —N(CH₃)₂, or Nsubstituted pyrrolidinyl and ii) administering said derivative to saidsubject. In further embodiments, nicotinoyl riboside and derivativesthat inhibit inosine 5′-monophosphate dehydrogenase.

In some embodiments, the invention relates to a method of managing,preventing or treating neurodegenerative diseases comprising i)providing a) a subject having symptoms of a neurodegenerative diseaseand b) a composition comprising a nicotinyl riboside derivativedisclosed herein and ii) administering said derivative to said subject.

In further embodiments, said composition comprises a compound having thefollowing formula.

wherein R is —OMe, —OEt, —OCH₂CH₂OH, —NHMe, —NHEt, —NHCH₂CH₂OH,—NHCH₂CH═CH₂, —NHCH(CH₃)₂, —NHCH₂CH₂NH₂, —NHcyclopropyl, —N(CH₃)₂, or Nsubstituted pyrrolidinyl.

In one aspect, the invention relates to the use of chemicalintermediates disclosed herein to make nicotinic acid riboside,nicotinoyl riboside, or nicotinamide riboside, or derivatives.

In another aspect, the invention relates the use of nicotinoyl ribosidesor derivatives for increasing the lifespan of a cell, and treatingand/or preventing a wide variety of diseases and disorders including,for example, diseases or disorders related to aging or stress, diabetes,obesity, neurodegenerative diseases, cardiovascular disease, bloodclotting disorders, inflammation, cancer, eye disorders, and/orflushing.

A method for treating or preventing one or more of cataracts,retinopathy, retinitis pigmentosa, ocular neuritis or a vascular diseaseof the capillary beds of the eye, comprising administering to a subjecta therapeutically effective amount of nicotinoyl ribosides orderivative.

A method for reducing the weight of a subject, or preventing weight gainin a subject, comprising administering to a subject in need thereof atherapeutically effective amount of a nicotinoyl ribosides orderivative.

A method for prolonging the lifespan of a subject comprisingadministering to a subject a therapeutically effective amount of anicotinoyl ribosides or derivative.

In further embodiments, the invention relates to a method of reducingdrug toxicity comprising: i) providing: a) a subject, b) a drug, c) acomposition comprising a nicotinyl riboside derivative; ii)administering said drug to said subject under conditions such that saiddrug produces an adverse reaction; and iii) administering saidcomposition under conditions such that said adverse reaction is reduced.In further embodiment, said drug is a statin. In further embodiment,said subject is a human In further embodiments, said compositioncomprises a compound having the following formula:

wherein R is —OMe, —OEt, —OCH₂CH₂OH, —NHMe, —NHEt, —NHCH₂CH₂OH,—NHCH₂CH═CH₂, —NHCH(CH₃)₂, —NHCH₂CH₂NH₂, —NHcyclopropyl, —N(CH₃)₂, or Nsubstituted pyrrolidinyl.

In further embodiment, the invention relates to a method of preventingdrug toxicity comprising: i) providing: a) a subject, b) a drug, c) acomposition comprising a nicotinamide riboside derivative; ii)administering said drug in a concentration that typically producestoxicity in said subject and said nicotinamide riboside composition tosaid subject under conditions such that drug toxicity is prevented.

In some embodiments, the invention relates to a method in which thehealth of a cell is managed comprising: i) providing a) a compositioncomprising a compound having the following formula:

and salt complexes thereof wherein, R¹ is amine, alkylamine, substitutedalkylamine, dialkylamino, substituted dialkylamino, alkyloxy,substituted alkyloxy, aryloxy, substituted aryloxy, arylthio,substituted arylthio, alkylthio, or substituted alkylthio; and R³, R⁴and R⁵ are the same or different and, at each occurrence hydrogen,independently acyl or substituted acyl; and b) cells in growth media;and ii) mixing said composition and said cells in growth media underconditions such that the heath of said cells is managed. In furtherembodiments, said cells are stem cells. In further embodiments, thecomposition increases the amount of NAD+ in the cells.

In some embodiments, the invention relates to a method of reducing anadverse drug reaction comprising: i) providing: a) a subject, b) a drug,and c) a composition comprising a nicotinoyl riboside or derivative; ii)administering said drug to said subject under conditions such that saiddrug produces an adverse reaction; and iii) administering saidcomposition under conditions such that said adverse reaction is reduced.In further embodiments, said drug is a statin. In further embodiments,said subject is a human.

In some embodiments, the invention relates to a method of preventing anadverse drug reaction comprising: i) providing: a) a subject, b) a drug,and c) a composition comprising a nicotinoyl riboside or derivative; ii)administering said drug in a concentration that typically produces anadverse drug reaction in said subject and iii) administering saidcomposition to said subject under conditions such that an adverse drugreaction is prevented. In further embodiments, said drug is a statin. Infurther embodiments, said subject is a human.

In some embodiments, the invention relates to using nicotinoyl ribosidesor derivatives for increasing cellular sensitivity to stress (includingincreasing radiosensitivity and/or chemosensitivity), increasing theamount and/or rate of apoptosis, treatment of cancer (optionally incombination another chemotherapeutic agent), stimulation of appetite,and/or stimulation of weight gain.

In some embodiments, the invention relates to a method of treatingcancer comprising i) providing a) a subject diagnosed with cancer and b)a nicotinoyl riboside or derivative; and ii) administering saidderivative to said subject under conditions such that said cancer isreduced. In additional embodiments, the invention relates to a method ofpreventing cancer comprising i) providing a) a subject at risk forcancer and b) a nicotinoyl riboside or derivative; and ii) administeringsaid derivative to said subject under conditions such that said canceris prevented. In further embodiments, said nicotinoyl riboside ornicotinamide derivatives inhibits inosine 5′-monophosphatedehydrogenase. In further embodiments, said nicotinoyl riboside ornicotinamide derivatives reduce the amount of NAD+ in the cellscomprising the cancer. In further embodiments, said subject is a human.In further embodiments, said cancer is selected from pancreatic cancer,endometrial cancer, small cell and non-small cell cancer of the lung(including squamous, adneocarcinoma and large cell types), squamous cellcancer of the head and neck, bladder, ovarian, cervical, breast, renal,CNS, and colon cancers, myeloid and lymphocyltic leukemia, lypmphoma,heptic tumors, medullary thyroid carcinoma, multiple myeloma, melanoma,retinoblastoma, and sarcomas of the soft tissue and bone.

In additional embodiments, the invention relates to a method of treatingor preventing neurodegenerative diseases neurodegenerative or conditioncomprising i) providing a) a subject diagnosed with, at risk for, orhaving symptoms of a neurodegenerative disease or neurodegenerativecondition and b) a composition comprising a nicotinoyl riboside orderivative; and ii) administering said derivative to said subject underconditions such that said neurodegenerative disease or neurodegenerativecondition is reduced. In further embodiments, said subject is a human.In further embodiments, said neurodegenerative disease orneurodegenerative condition is essential tremor, parkinson disease,alzheimer disease, huntington disease, ataxia, catatonia, epilepsy,neuroleptic malignant syndrome, dystonia, mental retardation,neuroacanthocytosis, Pelizaeus-Merzbacher, progressive supranuclearpalsy, Striatonigral degeneration, Tardive dyskinesias, damage followingstroke or trauma, or a lysosomal storage disorder, including lipidstorage disorders (including Gaucher's and Niemann-Pick diseases),gangliosidosis (including Tay-Sachs disease), leukodystrophies,mucopolysaccharidoses, glycoprotein storage disorders, andmucolipidoses.

In some embodiments, the invention relates to a method of increasingintracellular concentrations of NAD+ comprising: a) providing i) a celland ii) a nicotinoyl riboside or derivative; and b) mixing said cell andnicotinamide riboside or derivative under conditions such thatintracellular concentrations of NAD+ increase. In additionalembodiments, the invention relates to a method of decreasingintracellular concentrations of NAD+ comprising: a) providing i) a celland ii) a nicotinoyl riboside or derivative; and b) mixing said cell andnicotinamide riboside or derivative under conditions such thatintracellular concentrations of NAD+ decrease. In further embodimentssaid nicotinamide riboside is a compound having the following formula:

and salt complexes thereof wherein, R¹ is amine, alkylamine, substitutedalkylamine, dialkylamino, substituted dialkylamino, alkyloxy,substituted alkyloxy, aryloxy, substituted aryloxy, arylthio,substituted arylthio, alkylthio, or substituted alkylthio; and R³, R⁴and R⁵ are the same or different and, at each occurrence hydrogen,independently acyl or substituted acyl.

In some embodiments, the invention relates to a method of preventingdiabetes comprising i) providing a) a subject at risk for type I or typeII diabetes and b) a composition comprising a nicotinoyl riboside orderivative; and ii) administering said composition to said subject underconditions such that said diabetes is prevented. In further embodiments,said subject is a human. In further embodiments, said subject is obese.In additional embodiments, the invention relates to a method of treatingdiabetes comprising i) providing a) a subject diagnosed with or withsymptoms of type I or type II diabetes and b) a composition comprising anicotinoyl riboside or derivative; and ii) administering saidcomposition to said subject under conditions such that said symptoms arereduced. In further embodiments, said subject is a human. In furtherembodiments, said subject is obese.

In some embodiments, the invention relates to a method of preventing asubject from acquiring insulin insensitivity comprising i) providing a)a subject and b) a composition comprising a nicotinoyl riboside orderivative; and ii) administering said composition to said subject underconditions such that insulin insensitivity is prevented. In furtherembodiments, said subject is a human. In further embodiments, saidsubject is obese.

In other embodiments, the invention relates to a method of treating orpreventing insulin resistance disorders by administering a nicotinoylriboside or derivative disclosed herein.

In further embodiments, the invention relates to a method of preventinga subject from growing an abnormally large organ comprising i) providinga) a subject and b) a composition comprising a nicotinoyl riboside orderivative; and ii) administering said composition to said subject underconditions such that the growth of an abnormally large organ isprevented. In further embodiments, said subject is a human. In furtherembodiments, said organ is the liver. In further embodiments, saidsubject is obese.

In some embodiments, the invention relates to a method of increasing thenumber of cellular mitochondria in a subject comprising i) providing a)a subject and b) a composition comprising a nicotinoyl riboside orderivative; and ii) administering said composition to said subject underconditions such that the number of cellular mitochondria increase in anorgan of said subject increase. In further embodiments, said subject isa human. In further embodiments, said organ is the liver. In furtherembodiments, said subject is obese.

In further embodiments, the invention relates to a method of preventingheart disease in a subject comprising i) providing a) a subject and b) acomposition comprising a nicotinoyl riboside or derivative; and ii)administering said composition to said subject under conditions suchthat heart disease is decreased. In further embodiments, said subject isa human. In further embodiments, said subject is obese.

In another embodiment, the invention relates to a nicotinoyl riboside orderivative administered with one or more therapeutic agents for thetreatment or prevention of various diseases, including, for example,cancer, diabetes, neurodegenerative diseases, cardiovascular disease,blood clotting, inflammation, flushing, obesity, ageing, stress, forexample, administered with one or more of the following compounds:resveratrol, butein, fisetin, piceatannol, or quercetin. In an exemplaryembodiment, nicotinoyl riboside or derivative may be administered incombination with nicotinic acid. In another embodiment, nicotinoylriboside or derivative that decreases the level and/or activity of asirtuin protein may be administered with one or more of the followingcompounds: nicotinamide (NAM), suranim; NF023 (a G-protein antagonist);NF279 (a purinergic receptor antagonist); Trolox(6-hydroxy-2,5,7,8,tetramethylchroman-2-carboxylic acid);(−)-epigallocatechin (hydroxy on sites 3,5,7,3′,4′,5′);(−)-epigallocatechin gallate (Hydroxy sites 5,7,3′,4′,5′ and gallateester on 3); cyanidin chloride (3,5,7,3′,4′-pentahydroxyflavyliumchloride); delphinidin chloride (3,5,7,3′,4′,5′-hexahydroxyflavyliumchloride); myricetin (cannabiscetin; 3,5,7,3′,4′,5′-hexahydroxyflavone);3,7,3′,4′,5′-pentahydroxyflavone; gossypetin(3,5,7,8,3′,4′-hexahydroxyflavone), sirtinol; and splitomicin.

In further embodiments, the inventions relates to a nicotinoyl ribosideor derivative administered to subjects within the Kingdom Monera,including the true bacteria (eubacteria) and cyanobacteria (blue-greenalgae). Such administration may be to control the growth and/ormorphology of such subjects, for instance in the context ofbiotechnology production processes, or it may be in the context oftreatment of another subject or object that is infected with subjectswithin the Kingdom Monera.

In further embodiments, the invention relates to a nicotinoyl ribosideor derivative administered to subjects within the Kingdom Protista. Suchadministration may be in the context of treatment of another subject orobject that is infected with subjects within the Kingdom Protista.

In further embodiments, the invention relates to a nicotinoyl ribosideor derivative administered to subjects within the Kingdom Fungi. Suchadministration may be to control the growth and/or morphology of suchsubjects, for instance in the context of biotechnology productionprocesses, or it may be in the context of treatment of another subjector object that is infected with subjects within the Kingdom Fungi.

In further embodiments, the invention relates to a nicotinoyl ribosideor derivative administered to subjects within the Kingdom Plantae. Suchadministration may be to control the growth and/or morphology of suchsubjects, for instance in the context of biotechnology, agricultural, orhorticultural production processes. Such administration may be topromote the growth of such subjects, or to prevent the growth or killsuch subjects.

In further embodiments, the invention relates to a nicotinoyl ribosideor derivative marketed in compliance with the United States FederalFood, Drug, and Cosmetic Act (21 U.S.C. 321) as a dietary supplement.

In further embodiments, the invention relates to a nicotinoyl ribosideor derivative marketed in compliance with the United States FederalFood, Drug, and Cosmetic Act (21 U.S.C. 321) as a dietary supplementwith antioxidant properties.

In further embodiments, the invention relates to a nicotinoyl ribosideor derivative marketed in compliance with the United States FederalFood, Drug, and Cosmetic Act (21 U.S.C. 321) as a dietary supplementwith health claims.

In further embodiments, the invention relates to a nicotinoyl ribosideor derivative marketed in compliance with the United States FederalFood, Drug, and Cosmetic Act (21 U.S.C. 321) as a dietary supplementwith health claims for reducing the risk of or preventing a disease.

In further embodiments, the invention relates to a nicotinoyl ribosideor derivative marketed in compliance with the United States FederalFood, Drug, and Cosmetic Act (21 U.S.C. 321) as a dietary supplementwith health claims for reducing the risk of or preventing any of thediseases described herein.

In further embodiments, the invention relates to a nicotinoyl ribosideor derivative marketed in compliance with the United States FederalFood, Drug, and Cosmetic Act (21 U.S.C. 321) as a dietary supplementwith structure/function claims.

In further embodiments, the invention relates to methods to synthesize anicotinoyl riboside or derivative that is to be marketed as a dietarysupplement.

BRIEF DESCRIPTION OF THE FIGURE

FIG. 1 shows a scheme of preferred methods of for preparing preferredcompositions.

FIG. 2 shows data on the improvement of cell survival when grown on amedia supplemented with nicotinamide riboside (NAR) as provided inexample 7.

FIG. 3 shows data on the increase of intracellular levels ofnicotinamide adenine dinucleotide (NAD+) when embryonic stem cells areexposed to a culture containing NAR as provided in example 6.

FIG. 4 shows data on effects of NAR on cell survival as provided inexample 8.

FIG. 5 illustrates the preparation of N-alkyl nicotinamide ribosides.

FIG. 6 illustrates kinetically (−20° C.) or thermodynamically (4° C.)controlled reactions of O-alkyl nicotinate ribosides with alkyl amines.

FIG. 7 shows NAD+ biosynthetic pathway in mammals. Abbreviations: NA,nicotinic acid; Npt, nicotinic acid phosphoribosyltransferase; NaMN,nicotinic acid mononucleotide; Nmnat, nicotinamide mononucleotideadenylyltransferase; NAAD, nicotinic acid adenine dinucleotide; NAM,nicotinamide; NMN, nicotinamide mononucleotide; NR, nicotinamideriboside; Nrk, nicotinamide riboside kinase; ART, ADP-ribosyltransferase; PARP, poly-ADP-polymerases; Nampt, nicotinamidephosphoribosyltransferase also known as pre-B cell colony-enhancingfactor (PBEF).

FIG. 8 demonstrates reduction of statin (lovastatin) toxicity for mouseembryonic stem cells treated with nicotinamide riboside (500 μM) forperiods of 2, 4 and 7 days. Statin only treated controls are indicatedby minus symbols. The bars represent the concentration of statin thatleads to 50% decrease in cell number or MTT assay, which measures cellviability. The two far left columns in each day are measured by cellcounting, the two far right columns are measured by MTT. As shown, NRtreatment significantly reduces the toxicity of statin on cell viabilityas determined by an increase in the amount of statin required to killcells.

FIG. 9 shows a bar chart of NAD+ content (pmoL) in ES cells aftertreated with different compounds.

FIG. 10 shows the percentage of cell death after 48-hour treatment withlovastatin (800 nM and 5 μM). Mouse ES cells were treated in thepresence of nicotinate riboside derivatives NAR (500 μM), OENR (1 mM),TAENR (1 mM). Abbreviations: NAR: Nicotinic acid riboside, OENR: O-ethylnicotinamide riboside, TAENR: tri-O-acetyl O′-ethyl nicotinamideriboside. Control cells were not treated with a derivative. Cell deathpercentage is determined by the percentage of total viable cells in thestatin treated wells versus the untreated (no statin) controls. Theeffect of each additive to cell death is computed similarly, bycomparison to a no statin control in which the nicotinate ribosidederivative is also present. Cell counts in untreated controls (nostatin) for each experimental group were similar, indicative of a lackof toxicity of the compounds on cells.

FIG. 11 shows the chemical structure of compounds tested in FIG. 10.

FIG. 12 illustrates data from cell proliferation assay with DMNR (1 mM)and ANR (1 mM) treatment. DMNR: N-dimethyl nicotinamide riboside, ANR:N-allyl nicotinamide riboside.

FIG. 13 shows chemical structures of compounds tested in FIG. 12.

FIG. 14 shows ¹H-NMR of nicotinamide riboside

FIG. 15 shows ¹H-NMR of O-ethyl nicotinate riboside.

FIG. 16 shows ¹H-NMR of N-methyl nicotinamide riboside.

FIG. 17 shows ¹H-NMR of N-ethyl nicotinamide riboside.

FIG. 18 shows ¹H-NMR of N-allyl nicotinamide riboside.

FIG. 19 shows ¹H-NMR of N-ethanol nicotinamide riboside.

FIG. 20 shows ¹H-NMR of dimethyl nicotinamide riboside.

DETAILED DESCRIPTION OF THE INVENTION

The invention relates to compositions of nicotinoyl ribosides andnicotinamide riboside derivatives and their methods of use. In someembodiments, the invention relates to methods of making nicotinoylribosides. In some embodiments, the invention relates to pharmaceuticalcompositions and nutritional supplements containing a nicotinoylriboside. In further embodiments, the invention relates to methods ofusing nicotinoyl ribosides and nicotinamide riboside derivatives thatpromote the increase of intracellular levels of nicotinamide adeninedinucleotide (NAD+) in cells and tissues for improving cell and tissuesurvival.

A “nicotinoyl riboside” compound means a substituted or unsubstitutedcompound of the following formula:

Nicotinoyl riboside “salts” refers to salts which make up thecomposition which specifically include the nicotinoyl riboside salthaving the partial formula

and Z is a counter ion, including chloride, bromide, iodide, alkoxide,toluenesulfonate, methylsulfonate, sulfonate, phosphate, or carboxylate(such as benzoate, succinate, acetate, glycolate, maleate, malate,fumarate, citrate, tartrate, ascorbate, cinnamoate, mandeloate, anddiphenylacetate).

As used herein, the term “nicotinoyl riboside component” refers thatpart of a composition that contains all of the nicotinoyl ribosidemolecules in a given composition, including all conformational andstereometric forms. In preferred embodiments, a given compound (e.g.designated by a structure) makes up a large percentage (e.g. by numberof molecules and/or by weight) of the nicotinoyl riboside component. Forexample, a given nicotinoyl riboside molecule may be present in anaqueous composition at a level where 70% of all the nicotinoyl ribosidemolecules are of that given compound, while most of the compositionitself is composed of water.

“Acyl” means an —C(═O)alkyl or —C(═O)aryl group.

“Adverse drug reaction” means any response to a drug that is noxious andunintended and occurs in doses for prophylaxis, diagnosis, or therapyincluding side effects, toxicity, hypersensitivity, drug interactions,complications, or other idiosyncrasy. Side effects are often adversesymptom produced by a therapeutic serum level of drug produced by itspharmacological effect on unintended organ systems (e.g., blurred visionfrom anticholinergic antihistamine). A toxic side effect is an adversesymptom or other effect produced by an excessive or prolonged chemicalexposure to a drug (e.g., digitalis toxicity, liver toxicity).Hypersensitivities are immune-mediated adverse reactions (e.g.,anaphylaxis, allergy). Drug interactions are adverse effects arisingfrom interactions with other drugs, foods or disease states (e.g.,warfarin and erythromycin, cisapride and grapefruit, loperamide andClostridium difficile colitis). Complications are diseases caused by adrug (e.g., NSAID-induced gastric ulcer, estrogen-induced thrombosis).The adverse drug reaction may be mediated by known or unknown mechanisms(e.g., Agranulocytosis associated with chloramphenicol or clozapine).Such adverse drug reaction can be determined by subject observation,assay or animal model well-known in the art.

“Alkyl” means a straight chain or branched, noncyclic or cyclic,unsaturated or saturated aliphatic hydrocarbon containing from 1 to 10carbon atoms, while the term “lower alkyl” has the same meaning as alkylbut contains from 1 to 6 carbon atoms. The term “higher alkyl” has thesame meaning as alkyl but contains from 2 to 10 carbon atoms.Representative saturated straight chain alkyls include methyl, ethyl,n-propyl, n-butyl, n-pentyl, n-hexyl, n-septyl, n-octyl, n-nonyl, andthe like; while saturated branched alkyls include isopropyl, sec-butyl,isobutyl, tert-butyl, isopentyl, and the like. Representative saturatedcyclic alkyls include cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl,and the like; while unsaturated cyclic alkyls include cyclopentenyl andcyclohexenyl, and the like. Cyclic alkyls are also referred to herein asa “homocycles” or “homocyclic rings.” Unsaturated alkyls contain atleast one double or triple bond between adjacent carbon atoms (referredto as an “alkenyl” or “alkynyl”, respectively). Representative straightchain and branched alkenyls include ethylenyl, propylenyl, 1-butenyl,2-butenyl, isobutylenyl, 1-pentenyl, 2-pentenyl, 3-methyl-1-butenyl,2-methyl-2-butenyl, 2,3-dimethyl-2-butenyl, and the like; whilerepresentative straight chain and branched alkynyls include acetylenyl,propynyl, 1-butynyl, 2-butynyl, 1-pentynyl, 2-pentynyl,3-methyl-1-butynyl, and the like.

“Alkylamino” and “dialkylamino” mean one or two alkyl moiety attachedthrough a nitrogen bridge (i.e., —N-alkyl) such as methylamino,ethylamino, dimethylamino, diethylamino, and the like.

“Alkylthiol” means an alkyl moiety attached through a sulfur bridge(i.e., —S— alkyl).

“Alkyloxy” means an alkyl moiety attached through an oxygen bridge(i.e., —O— alkyl) such as methoxy, ethoxy, and the like.

“Alkoxide” means an alkyl moiety attached to a negatively charged oxygenatom (i.e., —Oalkyl) such as methoxide or ethoxide.

Within the context of certain embodiment, an “amine” means —NH₂ or“ammonia” the gas NH₃.

“Aryl” means an aromatic carbocyclic moiety such as phenyl or naphthyl.

“Aryloxy” means an aryl moiety attached through an oxygen bridge (i.e.,—O-aryl).

“Arylthio” means an alkyl moiety attached through a sulfur bridge (i.e.,—S-aryl).

“Aminooxy” means an amino moiety attached through an oxygen bridge(i.e., —O—NH₂).

“Hydroxylamino” means hydroxyl moiety attached through an amine bridge(i.e., —NH—OH).

“O-alkyloxyamino” means an alkyloxy moeity attached through an aminebridge (i.e., —NH—O-alkyl).

“N-alkylaminooxy” means an alkylamino moeity attached through an oxygenbridge (i.e., —O—NH-alkyl).

“Hydrazino” means an amino moiety attached through a nitrogen bridge(i.e., —NH—NH₂) depending on the context.

“Alkylhydrazino” means an alkyl moiety attached through a hydrazinebridge (i.e., —NH—NH-alkyl).

“Cancer” means any of various cellular diseases with malignant neoplasmscharacterized by the proliferation of anaplastic cells. It is notintended that the diseased cells must actually invade surrounding tissueand metastasize to new body sites. Cancer can involve any tissue of thebody and have many different forms in each body area. Most cancers arenamed for the type of cell or organ in which they start.

Within the context of certain embodiments, whether “cancer is reduced”may be identified by a variety of diagnostic manners known to one skillin the art including, but not limited to, observation the reduction insize or number of tumor masses or if an increase of apoptosis of cancercells observed, e.g., if more than a 5% increase in apoptosis of cancercells is observed for a sample compound compared to a control withoutthe compound. It may also be identified by a change in relevantbiomarker or gene expression profile, such as PSA for prostate cancer,her2 for breast cancer, or others. For example, reduction of cancer maybe identified in vitro using the following conditions for evaluation ofapoptosis: i) Jurkat human T-cell leukemia cells are passed into flasks(250 mL, 75 cm²) with 20 mL of supporting media; ii) after incubation at37° C. with 5% CO₂, sample compound (or absent control) is added to aflask to make final concentration at 1 mM, and cells are incubated foranother day; iii) cells are treated with 10 μM camptothecin andincubated with SYTOX Green reagent and annexin V allophycocyanin (APC)conjugate (invitrogen) and iv) Flow cytometry at 488 nm and 633 nmexcitation. In cells undergoing apoptosis, phosphatidylserine (PS) istransferred from the cytoplasmic surface of the cell membrane to theouter leaflet. Annexin V has a high affinity for PS and dye conjugatesprovide indication of apoptosis by phosphatidylserine exposure andmembrane integrity.

“Cells” means the structural unit of an organism consisting of one ormore nuclei, cytoplasm, and various organelles, all surrounded by asemipermeable cell membrane.

“Growth media” are compositions used to grow microorganisms or cells inculture. There are different sorts of media for growing different sortsof cells. The biggest difference in growth media are between those usedfor growing cells in culture (cell culture uses specific cell typesderived from plants or animals) and those used for growingmicroorganisms (usually bacteria or yeast). These differences arise dueto the fact that cells derived from whole organisms and grown in cultureare often incapable of growth without the provision of certainrequirements, such as hormones or growth factors which usually occur invivo. In the case of animal cells these requirements are often providedby the addition of blood serum to the medium. These media are often redor pink due to the inclusion of pH indicators. Growth media forembryonic stem cells preferably contains minimal essential medium, i.e.,Eagle's: amino acids, salts (Ferric nitrate nonahydrate, Potassiumchloride, Magnesium sulfate, Sodium chloride, Sodium dihydrogenphosphate), vitamins, (Ascorbic acid, Folic acid, Nicotinamide,Riboflavin, B-12) or Dulbecco's: additionally iron, glucose;non-essential amino acids, sodium pyruvate, β-mercaptoethanol,L-glutamine, fetal bovine serum and Leukemia Inhibitory Factor (LIF). Inthe case of microorganisms, there are no such limitations as they areoften single cell organisms. One other major difference is that animalcells in culture are often grown on a flat surface to which they attach,and the medium is provided in a liquid form, which covers the cells.Bacteria such as Escherichia coli (E. coli, the most commonly usedmicrobe in laboratories) may be grown on solid media or in liquid media,liquid nutrient medium is commonly called nutrient broth. The preferredgrowth media for microorganisms are nutrient broth or Luria-Bertanimedium (L-B medium). Bacteria grown in liquid cultures often formcolloidal suspensions. When agar (a substance which sets into a gel) isadded to a liquid medium it can be poured into petri dishes where itwill solidify (these are called agar plates) and provide a solid mediumon which microbes may be cultured.

“Heteroaryl” means an aromatic heterocycle ring of 5- to 10 members andhaving at least one heteroatom selected from nitrogen, oxygen andsulfur, and containing at least 1 carbon atom, including both mono- andbicyclic ring systems. Representative heteroaryls are furyl,benzofuranyl, thiophenyl, benzothiophenyl, pyrrolyl, indolyl,isoindolyl, azaindolyl, pyridyl, quinolinyl, isoquinolinyl, oxazolyl,isooxazolyl, benzoxazolyl, pyrazolyl, imidazolyl, benzimidazolyl,thiazolyl, benzothiazolyl, isothiazolyl, pyridazinyl, pyrimidinyl,pyrazinyl, triazinyl, cinnolinyl, phthalazinyl, and quinazolinyl.

“Heteroarylalkyl” means an alkyl having at least one alkyl hydrogen atomreplaced with a heteroaryl moiety, such as —CH₂pyridinyl,—CH₂pyrimidinyl, and the like.

“Heterocycle” (also referred to herein as a “heterocyclic ring”) means a4- to 7-membered monocyclic, or 7- to 10-membered bicyclic, heterocyclicring which is either saturated, unsaturated, or aromatic, and whichcontains from 1 to 4 heteroatoms independently selected from nitrogen,oxygen and sulfur, and wherein the nitrogen and sulfur heteroatoms maybe optionally oxidized, and the nitrogen heteroatom may be optionallyquaternized, including bicyclic rings in which any of the aboveheterocycles are fused to a benzene ring. The heterocycle may beattached via any heteroatom or carbon atom. Heterocycles includeheteroaryls as defined above. Thus, in addition to the heteroarylslisted above, heterocycles also include morpholinyl, pyrrolidinonyl,pyrrolidinyl, piperidinyl, hydantoinyl, valerolactamyl, oxiranyl,oxetanyl, tetrahydrofuranyl, tetrahydropyranyl, tetrahydropyridinyl,tetrahydroprimidinyl, tetrahydrothiophenyl, tetrahydrothiopyranyl,tetrahydropyrimidinyl, tetrahydrothiophenyl, tetrahydrothiopyranyl, andthe like.

“Heterocyclealkyl” means an alkyl having at least one alkyl hydrogenatom replaced with a heterocycle, such as —CH₂morpholinyl, and the like.

“Homocycle” (also referred to herein as “homocyclic ring”) means asaturated or unsaturated (but not aromatic) carbocyclic ring containingfrom 3-7 carbon atoms, such as cyclopropane, cyclobutane, cyclopentane,cyclohexane, cycloheptane, cyclohexene, and the like.

“Isomers” means any of two or more substances that are composed of thesame elements in the same proportions but differ in the threedimensional arrangement of atoms including enantiomeric (i.e., mirrorimages) and diastereomeric isomers.

“Methylene” means —CH₂—.

The term “derivative” when used in relation to a chemical compoundrefers to a similar structure that upon application, e.g.,administration to a subject, is capable of providing, directly orindirectly, the function said chemical compound is disclosed to have. Inthe context of certain embodiments, a “nicotinamide riboside (NAR)derivative” is understood to not include the compound NAR.

A “tumor” means an abnormal mass of tissue growth that may be classifiedas benign or malignant.

The term “manage” when used in connection with a disease or conditionmeans to provide beneficial effects to a subject being administered witha prophylactic or therapeutic agent, which does not result in a cure ofthe disease. In certain embodiments, a subject is administered with oneor more prophylactic or therapeutic agents to manage a disease so as toprevent the progression or worsening of the disease.

Within the context of certain embodiments, the “health of a cell ismanaged” if more than a 3% increase in cell survival is observed for asample compound compared to a control without the compound under thefollowing conditions: i) cells are passed into flasks (250 mL, 75 cm²)with 20 mL of media; ii) after 2 days incubation at 37° C. with 5% CO₂,sample compound (or absent control) is added to a flask to make finalconcentration at 1 mM, and cells are incubated for another day; iii) theagent methyl methanesulfonate (mms) (200 mM) is added to flask to makefinal concentration of 1 mM; iv) after 3 hrs incubation, the cells areharvested and the living ones are counted for the percentage of deathwith trypan blue (count cells on hemacytometer: dead cells stain blue,while live cells exclude trypan blue).

As used herein, the terms “prevent” and “preventing” include theprevention of the recurrence, spread or onset. It is not intended thatthe present invention be limited to complete prevention. In someembodiments, the onset is delayed, or the severity of the disease isreduced.

As used herein, the terms “purified isomer” and “purified isomercomposition” are meant to indicate a composition (e.g. derived from aracemic mixture or synthesized de novo) wherein one isomer has beenenriched (e.g., alpha-isomer) over the other (e.g., beta-isomer), andmore preferably, wherein the other isomer represents less than 10%, andmore preferably less than 7%, and still more preferably, less than 2% ofthe preparation.

Purified compositions in accordance with the invention preferablycontain less than 5% mass/mass (m/m), advantageously less than 3% m/m,of impurities. It is to be understood that references herein to“impurities” are to be understood as to include unwanted reactionproducts that are not isomers formed during synthesis and does notinclude residual solvents remaining from the process used in thepreparation of the composition or excipients used in pharmaceuticalpreparations.

The expression “essentially free” of a molecule means that the moleculeis present in a composition only as an unavoidable impurity.

“Subject” means any animal, preferably a human patient, livestock, ordomestic pet.

A “stem cell” is a cell that has the ability to divide (self replicate)and given the right signals, stem cells can give rise (differentiate) toseveral different cell types that make up a living organism. Many of theterms used to define stem cells depend on the behavior of the cells inthe intact organism (in vivo), under specific laboratory conditions (invitro), or after transplantation in vivo. For example, the fertilizedegg is said to be totipotent because it has the potential to generateall the cells and tissues that make up an embryo and that support itsdevelopment in utero. The fertilized egg divides and differentiatesuntil it produces a mature organism. Other cells, which are importantfor embryonic development but are not incorporated into the body of theembryo, include the extraembryonic tissues, placenta, and umbilicalcord. All of these cells are generated from a single, totipotentcell—the zygote, or fertilized egg.

“Pluripotent” cells have the potential to give rise to any type of cell,a property observed in the natural course of embryonic development.Unipotent stem cell, a term that is applied to a cell in adultorganisms, means that the cells in question are capable ofdifferentiating along only one lineage. The adult stem cells in manydifferentiated, undamaged tissues are typically unipotent and give riseto just one cell type under normal conditions. This process allows for asteady state of self-renewal for the tissue. However, if the tissuebecomes damaged and the replacement of multiple cell types is required,pluripotent stem cells may become activated to repair the damage.

The “embryonic stem cell” (ES) is defined by its origin—that is from oneof the earliest stages of the development of the embryo, called theblastocyst. Specifically, embryonic stem cells are derived from theinner cell mass of the blastocyst at a stage before it would implant inthe uterine wall. At this stage, the preimplantation embryo of the mouseis made up of 150 cells and consists of a sphere made up of an outerlayer of cells (the trophectoderm), a fluid-filled cavity (theblastocoel), and a cluster of cells on the interior (the inner cellmass). Pluripotent ES cells can give rise to differentiated cell typesthat are derived from all three primary germ layers of the embryo(endoderm, mesoderm, and ectoderm).

The “adult” stem cell is an undifferentiated (unspecialized) cell thatis found in a differentiated (specialized) tissue; it can renew itselfand become specialized to yield all of the specialized cell types of thetissue from which it originated. Adult stem cells are capable ofself-renewal for the lifetime of the organism. Sources of adult stemcells have been found in the bone marrow, blood stream, cornea andretina of the eye, the dental pulp of the tooth, liver, skin,gastrointestinal tract, and pancreas. Unlike embryonic stem cells, thereare no identifiable adult stem cells that are capable of forming allcells of the body. However, blood stem cells (derived from mesoderm) areable to generate a number of differentiated cells including bothskeletal muscle (also derived from mesoderm) and neurons (derived fromectoderm).

The term “substituted” as used herein means any of the above groups(i.e., alkyl, aryl, arylalkyl, heteroaryl, heteroarylalkyl, homocycle,heterocycle and/or heterocyclealkyl) wherein at least one hydrogen atomis replaced with a substituent. In the case of an oxo substituent(“═O”), two hydrogen atoms are replaced. When substituted, one or moreof the above groups are “substituents.” Substituents within the contextof this invention include halogen, deuterium, tritium, borono, hydroxy,oxo, cyano, nitro, amino, alkylamino, dialkylamino, alkyl, alkoxy,alkylthio, haloalkyl, aryl, arylalkyl, heteroaryl, heteroarylalkyl,heterocycle and heterocyclealkyl, as well as a saccharide, —NR_(a)R_(b),—NR_(a)C(═O)R_(b), —NR_(a)C(═O)NR_(a)NR_(b),—NR_(a)C(═O)OR_(b)—NR_(a)SO₂R_(b), —C(═O)R_(a), C(═O)OR_(a),—C(═O)NR_(a)R_(b), —OC(═O)NR_(a)R_(b), —OR_(a), —SR_(a), —SOR_(a),—S(═O)₂R_(a), —OS(═O)₂R_(a) and —S(═O)₂OR_(a). In addition, the abovesubstituents may be further substituted with one or more of the abovesubstituents, such that the substituent substituted alky, substitutedaryl, substituted arylalkyl, substituted heterocycle or substitutedheterocyclealkyl. R_(a) and R_(b) in this context may be the same ordifferent and independently hydrogen, alkyl, haloalkyl, substitutedalkyl, aryl, substituted aryl, arylalkyl, substituted arylalkyl,heterocycle, substituted heterocycle, heterocyclealkyl or substitutedheterocyclealkyl. In the context of certain embodiments, a compound maybe described as “unsubstituted” meaning that the compound does notcontain extra substituents attached to the compound. An unsubstitutedcompound refers to the chemical makeup of the compound without extrasubstituents, e.g., the compound does not contain protecting group(s).For example, unsubstituted proline is the proline amino acid even thoughthe amino group of proline may be considered disubstituted with alkylgroups.

As used herein, the terms “treat” and “treating” are not limited to thecase where the subject (e.g. patient) is cured and the disease iseradicated. Rather, the present invention also contemplates treatmentthat merely reduces symptoms, and/or delays disease progression.“Diabetes” refers to high blood sugar or ketoacidosis, as well aschronic, general metabolic abnormalities arising from a prolonged highblood sugar status or a decrease in glucose tolerance. “Diabetes”encompasses both the type I and type II (Non Insulin Dependent DiabetesMellitus or NIDDM) forms of the disease. The risk factors for diabetesmay include but are not limited to the following factors: waistline ofmore than 40 inches for men or 35 inches for women, blood pressure of130/85 mmHg or higher, triglycerides above 150 mg/dl, fasting bloodglucose greater than 100 mg/dl or high-density lipoprotein of less than40 mg/dl in men or 50 mg/dl in women.

The term “hyperinsulinemia” refers to a state in an individual in whichthe level of insulin in the blood is higher than normal.

The term “insulin resistance” refers to a state in which a normal amountof insulin produces a subnormal biologic response relative to thebiological response in a subject that does not have insulin resistance.

An “insulin resistance disorder,” as discussed herein, refers to anydisease or condition that is caused by or contributed to by insulinresistance. Examples include: diabetes, obesity, metabolic syndrome,insulin-resistance syndromes, syndrome X, insulin resistance, high bloodpressure, hypertension, high blood cholesterol, dyslipidemia,hyperlipidemia, dyslipidemia, atherosclerotic disease including stroke,coronary artery disease or myocardial infarction, hyperglycemia,hyperinsulinemia and/or hyperproinsulinemia, impaired glucose tolerance,delayed insulin release, diabetic complications, including coronaryheart disease, angina pectoris, congestive heart failure, stroke,cognitive functions in dementia, retinopathy, peripheral neuropathy,nephropathy, glomerulonephritis, glomerulosclerosis, nephrotic syndrome,hypertensive nephrosclerosis some types of cancer (such as endometrial,breast, prostate, and colon), complications of pregnancy, poor femalereproductive health (such as menstrual irregularities, infertility,irregular ovulation, polycystic ovarian syndrome (PCOS)), lipodystrophy,cholesterol related disorders, such as gallstones, cholescystitis andcholelithiasis, gout, obstructive sleep apnea and respiratory problems,osteoarthritis, and prevention and treatment of bone loss, e.g.osteoporosis.

“Obese” individuals or individuals suffering from obesity are generallyindividuals having a body mass index (BMI) of at least 25 or greater.Obesity may or may not be associated with insulin resistance.

The term “prophylactic” or “therapeutic” treatment is art-recognized andrefers to administration of a drug to a host. If it is administeredprior to clinical manifestation of the unwanted condition (e.g., diseaseor other unwanted state of the host animal) then the treatment isprophylactic, i.e., it protects the host against developing the unwantedcondition, whereas if administered after manifestation of the unwantedcondition, the treatment is therapeutic (i.e., it is intended todiminish, ameliorate or maintain the existing unwanted condition or sideeffects therefrom).

Nicotinamide Adenine Dinucleotide Pathway

Nicotinamide adenine dinucleotide (NAD or NAD+) is important as aco-enzyme for different enzymes. Recent studies depicted that being theco-substrate of SIR2 (silent information regulator 2), NAD+ has a rolein regulating multiple biological processes, such as p53 regulatedapoptosis, fat storage, stress resistance, and gene silencing. Withoutlimiting the potential uses of the compositions described herein by anysingle theory, there are various pathways through which NAR is currentlythought to be metabolized. Nicotinamide riboside (NAR) is known as aNAD+ precursor for both human and yeast. It is able to enter a salvagepathway that leads to biological synthesis of NAD+ under the action ofthe enzyme nicotinamide riboside kinase (Nrk). NAR is converted to NMNby Nrk, which is then converted to NAD+ by the enzyme nicotinamidemononucleotide adenylytransferase (Nmnat). (see FIG. 7). Alternatively,nicotinamide riboside can enter NAD metabolism by means of othermetabolic paths, which would include action from enzymes that separatethe nicotinamide moiety from the sugar. Such a path would include theaction of phosphorylases that have been shown to degrade NAR in cells toform nicotinamide and ribose-1-phosphate. Nicotinamide is competent toenter NAD+ metabolism and is converted to NAD+ by the action of theenzyme nicotinamide pyrophosphoribosyltransferase.

Sirtuins are class III histone deacetylases (HDACs) and are ADP-ribosyltransferases also. They deacetylate lysine residues in a novel chemicalreaction that consumes nicotinamide adenine dinucleotide (NAD+),releasing nicotinamide, O-acetyl-ADPribose (AADPR), and the deacetylatedsubstrate. Altering intracellular NAD+ levels can improve the health ofa cell, but introduction of compounds that enter NAD+ metabolic pathwayscan also prove toxic to cells. In some embodiments, the inventionrelates to the use of compounds disclosed herein to manipulate NAD+levels, to modulate the activity of sirtuins and other ADP-ribosyltransferases, and to modulate IMPHD. These embodiments are used todestroy or weaken the defenses of cancer cells, or to promote survivalof neurons, myocytes, or stem cells via addition to growth media.

Nicotinic acid is an effective agent in controlling low-densitylipoprotein cholesterol, increasing high-density lipoproteincholesterol, and reducing triglyceride and lipoprotein (a) levels inhumans. Though nicotinic acid treatment affects all of the key lipids inthe desirable direction and has been shown to reduce mortality in targetpopulations, its use is limited because of a side effect of heat andredness termed flushing. Further, nicotinamide protects against strokeinjury in model systems, presumably due to multiple mechanisms includingincreasing mitochondrial NAD+ levels.

Accordingly, one embodiment, of the invention relates to the use ofcompositions comprising compounds disclosed herein that work through thenicotinamide riboside kinase pathway or other pathways of NAD+biosynthesis which have nutritional and/or therapeutic value inimproving plasma lipid profiles, prevention of stroke, and/or prolonginglife and well-being. For example, nicotinamide riboside or nicotinicacid riboside or derivatives may be bioavailable but ultimatelyconvertible by metabolism to nicotinic acid or nicotinamide, therebyproviding the benefits of these compounds on lowering cholesterol orenhancing tissue protection from conditions of oxidative stress, such asis seen in stroke. This pathway of metabolism would predictably increaseNAD metabolism in cells, besides providing effects observed fornicotinamide or nicotinic acid in therapy. Another aspect of theinvention relates to a method for prevention or treating a disease orcondition associated with the nicotinamide riboside kinase pathway orother pathways of NAD+ biosynthesis (see FIG. 7) by administering acomposition comprising compounds disclosed herein. Diseases orconditions which typically have altered levels of NAD+ or precursorswhich can be prevented or treated by supplementing a diet or therapeutictreatment regime with a composition comprising compounds disclosedherein include, but are not limited to, lipid disorders, (e.g.,dyslipidemia, hypercholesterolaemia or hyperlipidemia), stroke, type Iand II diabetes, cardiovascular disease, and other physical problemsassociated with obesity.

In some embodiments, the invention relates to the use of compoundsdisclosed herein as agonist and antagonist of enzymes in the pathway ofNAD+ biosynthesis. In further embodiments, the NAR derivatives disclosedherein are agonist, i.e., stimulates activities normally stimulated bynaturally occurring substances, of one or more surtuins, preferablySIRT1 in humans or Sir2p in yeast. In further embodiments, the NARderivatives are antagonist of one or surtuins.

Although the applicant does not intend that the invention be limited byany particular mechanism, this is believed to be useful because SIRT1deacetylates transcription factors such as FOXOs, p53, and nuclearfactor Kappa B (NFKB), that provide stress resistance, apoptosis, andinflammatory responses improving organism survival. The sirtuin SIRT1upregulates stress-protective pathways by deacetylation of FOXOtranscription factors, leading to increased transcription of GADD45 (DNArepair) and MnSOD (reactive oxygen detoxification). SIRT1 concomitantlydownregulates FOXO transcription of the proapoptotic factors Fas andBcl-2 interacting mediator of cell death. (BIM).

For example, the p53 network in normal, non-activated situations isnon-functional, but is activated in cells as a response to varioussignals by inhibiting the abnormal growth of cells and triggering ofprogrammed cell death. SIRT1 interacts with p53 and deacetylates theC-terminal regulatory domain. This activity downregulates effects of p53transcriptional activation on target genes.

Neurodegenerative Diseases

Axon degeneration occurs frequently in neurodegenerative diseases andperipheral neuropathies. The degeneration of transected axons is delayedin Wallerian degeneration slow (Wlds) mice with the overexpression of afusion protein with the nicotinamide adenine dinucleotide (NAD+)synthetic enzyme, nicotinamide mononucleotide adenylyltransferase(Nmnat1). Both Wld(s) and Nmnat1 themselves are functional in preventingaxon degeneration in neuronal cultures.

NAD+ levels decrease in injured, diseased, or degenerating neural cellsand preventing this NAD+ decline efficiently protects neural cells fromcell death. Araki & Milbrandt “Increased nuclear NAD+ biosynthesis andSIRT1 activation prevent axonal degeneration” Science. 2004 Aug. 13;305(5686):1010-3 and Wang et al., “A local mechanism mediatesNAD-dependent protection of axon degeneration” J Cell Biol.170(3):349-55 (2005) hereby incorporated by reference. As a number ofnicotinoyl riboside compounds disclosed herein are capable of increasingintracellular levels of NAD+, these compounds are useful as atherapeutic or nutritional supplement in managing injuries, diseases,and disorders effecting the central nervous system and the peripheralnervous system, including but not limited to trauma or injury to neuralcells, diseases or conditions that harm neural cells, andneurodegenerative diseases or syndromes. Some neurodegenerativediseases, neurodegenerative syndromes, diseases and conditions that harmneural cells, and injury to neural cells are described below.

Essential tremor (ET) is the most common movement disorder. It is asyndrome characterized by a slowly progressive postural and/or kinetictremor, usually affecting both upper extremities.

Parkinson disease (PD) is a progressive neurodegenerative disorderassociated with a loss of dopaminergic nigrostriatal neurons.

Alzheimer disease (AD) is the most common form of dementia. It is aprogressive degenerative disease of the brain, strongly associated withadvanced age. Over time, people with the disease lose their ability tothink and reason clearly, judge situations, solve problems, concentrate,remember useful information, take care of themselves, and even speak. Anumber of neurodegenerative disease such as Alzheimer's disease executetheir biological impact in the brain. It is preferred that nicotinoylriboside compounds disclosed herein are capable of passing theblood-brain-barrier (BBB).

Huntington disease (HD) is an incurable, adult-onset, autosomal dominantinherited disorder associated with cell loss within a specific subset ofneurons in the basal ganglia and cortex.

Ataxia is defined as an inability to maintain normal posture andsmoothness of movement. Neurologic symptoms and signs such as seizuresand movement disorders (eg, dystonia, chorea) may accompany ataxia.

Catatonia is a state of apparent unresponsiveness to external stimuli ina person who is apparently awake. Epilepsy is defined as a chroniccondition characterized by spontaneous, recurrent seizures; seizure isdefined as a clinical event associated with a transient,hypersynchronous neuronal discharge.

Neuroleptic malignant syndrome (NMS) refers to the combination ofhyperthermia, rigidity, and autonomic dysregulation that can occur as aserious complication of the use of antipsychotic drugs.

Chorea is an involuntary abnormal movement, characterized by abrupt,brief, nonrhythmic, nonrepetitive movement of any limb, often associatedwith nonpatterned facial grimaces. Chorea gravidarum (CG) is the termgiven to chorea occurring during pregnancy.

Cortical basal ganglionic degeneration (CBGD) clinical characteristicinclude progressive dementia, parkinsonism, and limb apraxia.Dysfunction of the central or peripheral nervous system pathways maycause autonomic dysfunction.

Dystonia is a syndrome of sustained muscle contractions, usuallyproducing twisting and repetitive movements or abnormal postures.Writer's cramp is a form of task-specific focal dystonia.

Mental retardation (MR) is a condition in which intellectual capacity islimited significantly. Developmental disability describes a conditionthat limits an individual's ability to perform activities and roles asexpected in a certain social environment. Frequently, MR anddevelopmental disabilities are present simultaneously as a consequenceof brain damage.

Neuroacanthocytosis is a progressive neurologic disease characterized bymovement disorders, personality changes, cognitive deterioration, axonalneuropathy, and seizures. Most patients have acanthocytosis onperipheral blood smear at some point during the course of the disease.

Pelizaeus-Merzbacher disease (PMD) and X-linked spastic paraplegia type2 (SPG2) are at opposite ends of a clinical spectrum of X-linkeddiseases caused by mutations of the same gene, the proteolipid protein 1(PLPI) gene, and resulting in defective central nervous system (CNS)myelination. Clinical signs usually include some combination ofnystagmus, stridor, spastic quadriparesis, hypotonia, cognitiveimpairment, ataxia, tremor, and diffuse leukoencephalopathy on MRIscans.

Progressive supranuclear palsy (PSP), also known asSteele-Richardson-Olszewski syndrome, is a neurodegenerative diseasethat affects cognition, eye movements, and posture.

Striatonigral degeneration (SND) is a neurodegenerative disease thatrepresents a manifestation of multiple system atrophy (MSA). The othermanifestations are Shy-Drager syndrome (eg, autonomic failurepredominates) and sporadic olivopontocerebellar degeneration (sOPCA,cerebellum predominates).

Tardive dyskinesias (TDs) are involuntary movements of the tongue, lips,face, trunk, and extremities that occur in patients treated withlong-term dopaminergic antagonist medications. Although they areassociated with the use of neuroleptics, TDs apparently existed beforethe development of neuroleptics. People with schizophrenia appearespecially vulnerable to developing TDs after exposure to neuroleptics,toxins, and other agents. TDs are most common in patients withschizophrenia, schizoaffective disorder, or bipolar disorder who havebeen treated with antipsychotic medication for long periods, but TDsoccasionally occur in other patients as well.

The lysosomal storage diseases are a group of which over forty disordersare currently known that result from defects in lysosomal function.Lysosomes are cytoplasmic organelles that contain enzymes (specifically,acid hydrolases) that break macromolecules down to peptides, aminoacids, monosaccharides, nucleic acids and fatty acids. The lysosomalstorage diseases are classified by the nature of the primary storedmaterial involved, and can be broadly broken into the following: (ICD-10codes are provided where available)

-   -   (E75) lipid storage disorders (including Gaucher's and        Niemann-Pick diseases)    -   (E75.0-E75.1) gangliosidosis (including Tay-Sachs disease)    -   (E75.2) leukodystrophies    -   (E76.0) mucopolysaccharidoses    -   (E77) glycoprotein storage disorders    -   (E77.0-E77.1) mucolipidoses

Ischemic stroke occurs due to a loss of blood supply to part of thebrain, initiating the ischemic cascade. Brain tissue ceases to functionif deprived of oxygen for more than 60 to 90 seconds and after a fewhours will suffer irreversible injury possibly leading to death of thetissue, i.e., infarction. Atherosclerosis may disrupt the blood supplyby narrowing the lumen of blood vessels leading to a reduction of bloodflow, by causing the formation of blood clots within the vessel, or byreleasing showers of small emboli through the disintegration ofatherosclerotic plaques. Embolic infarction occurs when emboli formedelsewhere in the circulatory system, typically in the heart as aconsequence of atria fibrillation, or in the carotid arteries. Thesebreak off, enter the cerebral circulation, then lodge in and occludebrain blood vessels.

Due to collateral circulation, within the region of brain tissueaffected by ischemia there is a spectrum of severity. Thus, part of thetissue may immediately die while other parts may only be injured andcould potentially recover. The ischemia area where tissue might recoveris referred to as the ischemic penumbra.

As oxygen or glucose becomes depleted in ischemic brain tissue, theproduction of high energy phosphate compounds such as adeninetriphosphate (ATP) fails leading to failure of energy dependentprocesses necessary for tissue cell survival. This sets off a series ofinterrelated events that result in cellular injury and death. Theseinclude the failure of mitochondria, which can lead further towardenergy depletion and may trigger cell death due to apoptosis. Otherprocesses include the loss of membrane ion pump function leading toelectrolyte imbalances in brain cells. There is also the release ofexcitatory neurotransmitters, which have toxic effects in excessiveconcentrations.

Ischaemia also induces production of oxygen free radicals and otherreactive oxygen species. These react with and damage a number ofcellular and extracellular elements. Damage to the blood vessel liningor endothelium is particularly important. In fact, many antioxidantneuroprotectants such as uric acid and NXY-059 work at the level of theendothelium and not in the brain per se. Free radicals also directlyinitiate elements of the apoptosis cascade by means of redox signaling.

These processes are the same for any type of ischemic tissue and arereferred to collectively as the ischemic cascade. However, brain tissueis especially vulnerable to ischemia since it has little respiratoryreserve and is completely dependent on aerobic metabolism, unlike mostother organs.

Spinal cord injury, or myelopathy, is a disturbance of the spinal cordthat results in loss of sensation and mobility. The two common types ofspinal cord injury are:

-   -   Trauma: automobile accidents, falls, gunshots, diving accidents,        etc.    -   Disease: polio, spina bifida, tumors, Friedreich's ataxia, etc.

It is important to note that the spinal cord does not have to becompletely severed for there to be a loss of function. In fact, thespinal cord remains intact in most cases of spinal cord injury.

About 450,000 people in the United States live with spinal cord injury,and there are about 11,000 new spinal cord injuries every year. Themajority of them (78%) involve males between the ages of 16-30 andresult from motor vehicle accidents (42%), violence (24%), or falls(22%).

In a complete injury, there is no function below the level of theinjury. Voluntary movement is impossible and physical sensation isimpossible. Complete injuries are always bilateral, that is, both sidesof the body are affected equally.

A person with an incomplete injury retains some sensation below thelevel of the injury. Incomplete injuries are variable, and a person withsuch an injury may be able to move one limb more than another, may beable to feel parts of the body that cannot be moved, or may have morefunctioning on one side of the body than the other.

In addition to a loss of sensation and motor function below the point ofinjury, individuals with spinal cord injuries will often experienceother changes. Bowel and bladder function is associated with the sacralregion of the spine, so it is very common to experience dysfunction ofthe bowel and bladder. Sexual function is also associated with thesacral region, and is also affected very often. Injuries very high onthe spinal cord (C-1, C-2) will often result in a loss of manyinvoluntary functions, such as breathing, necessitating mechanicalventilators or phrenic nerve pacing. Other effects of spinal cord injurycan include an inability to regulate heart rate (and therefore bloodpressure), reduced control of body temperature, inability to sweat belowthe level of injury, and chronic pain. Physical therapy and orthopedicinstruments (e.g., wheelchairs, standing frames) are often necessary,depending on the location of the injury. Between about three weeks andtwelve years after lesions above T10 autonomic dysreflexia may occur.The injury may be located anywhere along the spinal cord; injuries areusually classified by location: Cervical injuries; Thoracic injuries;Lumbar and Sacral injuries; and Central Cord Syndrome. Central CordSyndrome is associated with ischemia, hemorrhage, or necrosis involvingthe central portions of the spinal cord (the large nerve fibers thatcarry information directly from the cerebral cortex). Corticospinalfibers destined for the legs are spared due to their more externallocation in the spinal cord.

Traumatic brain injury (TBI), traumatic injuries to the brain, alsocalled intracranial injury, or simply head injury, occurs when a suddentrauma causes brain damage. TBI can result from a closed head injury ora penetrating head injury and is one of two subsets of acquired braininjury (ABI). The other subset is non-traumatic brain injury (i.e.stroke, meningitis, anoxia). Parts of the brain that can be damagedinclude the cerebral hemispheres, cerebellum, and brain stem. Symptomsof a TBI can be mild, moderate, or severe, depending on the extent ofthe damage to the brain. Outcome can be anything from complete recoveryto permanent disability or death. A coma can also affect a child'sbrain. The damage from TBI can be focal, confined to one area of thebrain, or diffuse, involving more than one area of the brain. Diffusetrauma to the brain is frequently associated with concussion (a shakingof the brain in response to sudden motion of the head), diffuse axonalinjury, or coma. Localized injuries may be associated withneurobehavioral manifestations, hemiparesis or other focal neurologicdeficits.

Types of focal brain injury include bruising of brain tissue called acontusion and intracranial hemorrhage or hematoma, heavy bleeding in theskull. Hemorrhage, due to rupture of a blood vessel in the head, can beextra-axial, meaning it occurs within the skull but outside of thebrain, or intra-axial, occurring within the brain. Extra-axialhemorrhages can be further divided into subdural hematoma, epiduralhematoma, and subarachnoid hemorrhage. An epidural hematoma involvesbleeding into the area between the skull and the dura. With a subduralhematoma, bleeding is confined to the area between the dura and thearachnoid membrane. A subarachnoid hemorrhage involves bleeding into thespace between the surface of the brain and the arachnoid membrane thatlies just above the surface of the brain, usually resulting from a tearin a blood vessel on the surface of the brain. Bleeding within the brainitself is called an intracerebral hematoma. Intra-axial bleeds arefurther divided into intraparenchymal hemorrhage which occurs within thebrain tissue itself and intraventricular hemorrhage which occurs intothe ventricular system.

TBI can result from a closed head injury or a penetrating head injury. Aclosed injury occurs when the head suddenly and violently hits an objectbut the object does not break through the skull. A penetrating injuryoccurs when an object pierces the skull and enters brain tissue.

As the first line of defense, the skull is particularly vulnerable toinjury. Skull fractures occur when the bone of the skull cracks orbreaks. A depressed skull fracture occurs when pieces of the brokenskull press into the tissue of the brain. A penetrating skull fractureoccurs when something pierces the skull, such as a bullet, leaving adistinct and localized traumatic injury to brain tissue. Skull fracturescan cause cerebral contusion.

Another insult to the brain that can cause injury is anoxia. Anoxia is acondition in which there is an absence of oxygen supply to an organ'stissues, even if there is adequate blood flow to the tissue. Hypoxiarefers to a decrease in oxygen supply rather than a complete absence ofoxygen, and ischemia is inadequate blood supply, as is seen in cases inwhich the brain swells. In any of these cases, without adequate oxygen,a biochemical cascade called the ischemic cascade is unleashed, and thecells of the brain can die within several minutes. This type of injuryis often seen in near-drowning victims, in heart attack patients(particularly those who have suffered a cardiac arrest, or in people whosuffer significant blood loss from other injuries that then causes adecrease in blood flow to the brain due to circulatory (hypovolemic)shock.

Sometimes, health complications occur in the period immediatelyfollowing a TBI. These complications are not types of TBI, but aredistinct medical problems that arise as a result of the injury. Althoughcomplications are rare, the risk increases with the severity of thetrauma. Complications of TBI include immediate seizures, hydrocephalusor post-traumatic ventricular enlargement, cerebrospinal fluid leaks,infections, vascular injuries, cranial nerve injuries, pain, bed sores,multiple organ system failure in unconscious patients, and polytrauma(trauma to other parts of the body in addition to the brain).

About 25% of patients with brain contusions or hematomas and about 50%of patients with penetrating head injuries will develop immediateseizures, seizures that occur within the first 24 hours of the injury.These immediate seizures increase the risk of early seizures—defined asseizures occurring within 1 week after injury—but do not seem to belinked to the development of post-traumatic epilepsy (recurrent seizuresoccurring more than 1 week after the initial trauma). Generally, medicalprofessionals use anticonvulsant medications to treat seizures in TBIpatients only if the seizures persist.

Hydrocephalus or post-traumatic ventricular enlargement occurs whencerebrospinal fluid (CSF) accumulates in the brain resulting in dilationof the cerebral ventricles (cavities in the brain filled with CSF) andan increase in ICP. This condition can develop during the acute stage ofTBI or may not appear until later. Generally it occurs within the firstyear of the injury and is characterized by worsening neurologicaloutcome, impaired consciousness, behavioral changes, ataxia (lack ofcoordination or balance), incontinence, or signs of elevated ICP. Thecondition may develop as a result of meningitis, subarachnoidhemorrhage, intracranial hematoma, or other injuries. Treatment includesshunting and draining of CSF as well as any other appropriate treatmentfor the root cause of the condition.

Skull fractures can tear the membranes that cover the brain, leading toCSF leaks. A tear between the dura and the arachnoid membranes, called aCSF fistula, can cause CSF to leak out of the subarachnoid space intothe subdural space; this is called a subdural hygroma. CSF can also leakfrom the nose and the ear. These tears that let CSF out of the braincavity can also allow air and bacteria into the cavity, possibly causinginfections such as meningitis. Pneumocephalus occurs when air enters theintracranial cavity and becomes trapped in the subarachnoid space.

Infections within the intracranial cavity are a dangerous complicationof TBI. They may occur outside of the dura mater, below the dura, belowthe arachnoid (meningitis), or within the brain itself (abscess). Mostof these injuries develop within a few weeks of the initial trauma andresult from skull fractures or penetrating injuries. Standard treatmentinvolves antibiotics and sometimes surgery to remove the infectedtissue. Meningitis may be especially dangerous, with the potential tospread to the rest of the brain and nervous system.

Any damage to the head or brain usually results in some damage to thevascular system, which provides blood to the cells of the brain. Thebody's immune system can repair damage to small blood vessels, butdamage to larger vessels can result in serious complications. Damage toone of the major arteries leading to the brain can cause a stroke,either through bleeding from the artery (hemorrhagic stroke) or throughthe formation of a clot at the site of injury, called a thrombus orthrombosis, blocking blood flow to the brain (ischemic stroke). Bloodclots also can develop in other parts of the head. Symptoms such asheadache, vomiting, seizures, paralysis on one side of the body, andsemiconsciousness developing within several days of a head injury may becaused by a blood clot that forms in the tissue of one of the sinuses,or cavities, adjacent to the brain. Thrombotic-ischemic strokes aretreated with anticoagulants, while surgery is the preferred treatmentfor hemorrhagic stroke. Other types of vascular injuries includevasospasm and the formation of aneurysms.

Skull fractures, especially at the base of the skull, can cause cranialnerve injuries that result in compressive cranial neuropathies. All butthree of the 12 cranial nerves project out from the brainstem to thehead and face. The seventh cranial nerve, called the facial nerve, isthe most commonly injured cranial nerve in TBI and damage to it canresult in paralysis of facial muscles.

Pain, especially headache, is commonly a significant complication forconscious patients in the period immediately following a TBI. Seriouscomplications for patients who are unconscious, in a coma, or in avegetative state include bed or pressure sores of the skin, recurrentbladder infections, pneumonia or other life-threatening infections, andprogressive multiple organ failure.

Drug Toxicity

In some embodiments, the invention relates to the use of a nicotinoylriboside or derivative to prevent adverse effects and protect cells fromtoxicity. Toxicity may be an adverse effect of radiation or externalchemicals on the cells of the body. Examples of toxins arepharmaceuticals, drugs of abuse, and radiation, such as UV or X-raylight. Both radiative and chemical toxins have the potential to damagebiological molecules such as DNA. This damage typically occurs bychemical reaction of the exogenous agent or its metabolites withbiological molecules, or indirectly through stimulated production ofreactive oxygen species (eg, superoxide, peroxides, hydroxyl radicals).Repair systems in the cell excise and repair damage caused by toxins.

Enzymes that use NAD+ play an part in the DNA repair process.Specifically, the poly(ADP-ribose) polymerases (PARPs), particularlyPARP-1, are activated by DNA strand breaks and affect DNA repair. ThePARPs consume NAD+ as an adenosine diphosphate ribose (ADPR) donor andsynthesize poly(ADP-ribose) onto nuclear proteins such as histones andPARP itself. Although PARP activities facilitate DNA repair,overactivation of PARP can cause significant depletion of cellular NAD+,leading to cellular necrosis. The apparent sensitivity of NAD+metabolism to genotoxicity has led to pharmacological investigationsinto the inhibition of PARP as a means to improve cell survival.Numerous reports have shown that PARP inhibition increases NAD+concentrations in cells subject to genotoxicity, with a resultingdecrease in cellular necrosis. Nevertheless, cell death from toxicitystill occurs, presumably because cells are able to complete apoptoticpathways that are activated by genotoxicity. Thus, significant celldeath is still a consequence of DNA/macromolecule damage, even withinhibition of PARP. This consequence suggests that improvement of NAD+metabolism in genotoxicity can be partially effective in improving cellsurvival but that other players that modulate apoptotic sensitivity,such as sirtuins, may also play important roles in cell responses togenotoxins.

Physiological and biochemical mechanisms that determine the effects ofchemical and radiation toxicity in tissues are complex, and evidenceindicates that NAD+ metabolism is an important player in cell stressresponse pathways. For example, upregulation of NAD+ metabolism, vianicotinamide/nicotinic acid mononucleotide (NMNAT) overexpression, hasbeen shown to protect against neuron axonal degeneration, andnicotinamide used pharmacologically has been recently shown to provideneuron protection in a model of fetal alcohol syndrome and fetalischemia. Such protective effects could be attributable to upregulatedNAD+ biosynthesis, which increases the available NAD+ pool subject todepletion during genotoxic stress. This depletion of NAD+ is mediated byPARP enzymes, which are activated by DNA damage and can deplete cellularNAD+, leading to necrotic death. Another mechanism of enhanced cellprotection that could act in concert with upregulated NAD+ biosynthesisis the activation of cell protection transcriptional programs regulatedby sirtuin enzymes.

Examples of cell and tissue protection linked to NAD+ and sirtuinsinclude the finding that SIRT1 is required for neuroprotectionassociated with trauma and genotoxicity. SIRT1 can also decreasemicroglia-dependent toxicity of amyloid-beta through reduced NFKBsignaling. SIRT1 and increased NAD+ concentrations provideneuroprotection in a model of Alzheimer's disease. Sirtuins areNAD+-dependent enzymes that have protein deacetylase andADP-ribosyltransferase activities that upregulate stress responsepathways. Evidence indicates that SIRT1 is upregulated by calorierestriction and in humans could provide cells with protection againstapoptosis via downregulation of p53 and Ku70 functions. In addition,SIRT1 upregulates FOXO-dependent transcription of proteins involved inreactive oxygen species (ROS) detoxification, such as MnSOD. The sirtuinSIRT6 has been shown to participate in DNA repair pathways and to helpmaintain genome stability.

It seems likely that pharmacological agents that target both NAD+metabolism and sirtuins could provide tools to elucidate the involvementof these factors in modulating toxicity-induced tissue damage. Moreover,therapeutic options for treatment of acute and chronictissue-degenerative conditions could emerge if sirtuins and NAD+metabolism can be validated as providing enhanced tissue protection.Agents such as the plant polyphenols (eg, resveratrol), the niacinvitamins, and the compound nicotinamide riboside may enhance cellsurvival outcomes by increasing NAD+ biosynthesis, reducing NAD+depletion, and/or activating sirtuin enzymes.

Statins, more mechanistically known as 3-hydroxy-3-methyglutarylcoenzyme A reductase inhibitors (or HMG-CoA inhibitors) are some of theworld's most widely prescribed drugs. While statins are well toleratedat therapeutic doses, at higher doses and often in combination withother hypolipidaemic agents some potentially severe adverse effects havearisen. Most notably, cerivastatin (Baycol) was removed from the marketin 2000 after 31 deaths in the United States from drug-associatedrhabdomyolysis (breakdown of muscle fibers resulting in the release ofmuscle fiber contents into the circulation; some of these are toxic tothe kidney) and associated acute renal failure in patients takingcerivastatin. Statins are also known to have severe interactions withfibric acid derivatives, especially with gemfibrozil. Of the 31 peoplewho died taking cerivastatin, 12 were also taking gemfibrozil.

The most serious adverse effects of statins appear to occur in liver andmuscle cells, although it could be predicted that because of theirlipophilicity, cerebral effects might also be seen in some patients.

The exact mechanism of statin toxicities is unknown. The fact thattoxicities are dose-dependent makes plausible the hypothesis thattoxicities result from exaggeration of the drug's intended effect: inother words, cells die from lack of the downstream products of HMG-CoA.

HMG-CoA is the rate limiting enzyme in the mevalonate pathway, which,through three branches, leads to the synthesis of cholesterol, dolichol(the precursor to dolichol pyrophosphate, which is the first thing addedto proteins in post-translational glycosylation), and to ubiquinone,also known as Coenzyme Q (found in the membranes of endoplasmicreticulum, peroxisomes, lysosomes, vesicles and notably the innermembrane of the mitochondrion where it is an important part of theelectron transport chain; it is also has important antioxidantactivities).

However, it is likely that depletion of CoQ leads to a breakdown in theelectron transport chain, leading in turn to a buildup in NADH, and adepletion in NAD+. Further, the reduced form of CoQ10, CoQ10H2, has animportant cellular antioxidant function, which is to protect membranesand plasma lipoproteins against free radical-induced oxidation.

Aging/Stress

In one embodiment, the invention provides a method extending thelifespan of a cell, extending the proliferative capacity of a cell,slowing aging of a cell, promoting the survival of a cell, delayingcellular senescence in a cell, mimicking the effects of calorierestriction, increasing the resistance of a cell to stress, orpreventing apoptosis of a cell, by contacting the cell with nicotinoylriboside or derivative compound. In an exemplary embodiment, the methodscomprise contacting the cell with a nicotinoyl riboside or derivative.

The methods described herein may be used to increase the amount of timethat cells, particularly primary cells (i.e., cells obtained from anorganism, e.g., a human), may be kept alive in a cell culture. Embryonicstem (ES) cells and pluripotent cells, and cells differentiatedtherefrom, may also be treated with nicotinoyl riboside or derivativecompound to keep the cells, or progeny thereof, in culture for longerperiods of time. Such cells can also be used for transplantation into asubject, e.g., after ex vivo modification.

In one embodiment, cells that are intended to be preserved for longperiods of time may be treated with a nicotinoyl riboside or derivativecompound. The cells may be in suspension (e.g., blood cells, serum,biological growth media, etc.) or in tissues or organs. For example,blood collected from an individual for purposes of transfusion may betreated with a nicotinoyl riboside or derivative compound to preservethe blood cells for longer periods of time. Additionally, blood to beused for forensic purposes may also be preserved using a nicotinoylriboside or derivative compound. Other cells that may be treated toextend their lifespan or protect against apoptosis include cells forconsumption, e.g., cells from non-human mammals (such as meat) or plantcells (such as vegetables).

Nicotinoyl riboside or derivative compounds may also be applied duringdevelopmental and growth phases in mammals, plants, insects ormicroorganisms, in order to, e.g., alter, retard or accelerate thedevelopmental and/or growth process.

In another embodiment, a nicotinoyl riboside or derivative compounds maybe used to treat cells useful for transplantation or cell therapy,including, for example, solid tissue grafts, organ transplants, cellsuspensions, stem cells, bone marrow cells, etc. The cells or tissue maybe an autograft, an allograft, a syngraft or a xenograft. The cells ortissue may be treated with the a nicotinoyl riboside or derivativecompound prior to administration/implantation, concurrently withadministration/implantation, and/or post administration/implantationinto a subject. The cells or tissue may be treated prior to removal ofthe cells from the donor individual, ex vivo after removal of the cellsor tissue from the donor individual, or post implantation into therecipient. For example, the donor or recipient individual may be treatedsystemically with a nicotinoyl riboside or derivative compound or mayhave a subset of cells/tissue treated locally with a nicotinoyl ribosideor derivative compound. In certain embodiments, the cells or tissue (ordonor/recipient individuals) may additionally be treated with anothertherapeutic agent useful for prolonging graft survival, such as, forexample, an immunosuppressive agent, a cytokine, an angiogenic factor,etc.

In yet other embodiments, cells may be treated with a nicotinoylriboside or derivative compound that increases the level of NAD+ invivo, e.g., to increase their lifespan or prevent apoptosis. Forexample, skin can be protected from aging (e.g., developing wrinkles,loss of elasticity, etc.) by treating skin or epithelial cells with anicotinoyl riboside or derivative compound that increases the levelintracellular NAD+. In an exemplary embodiment, skin is contacted with apharmaceutical or cosmetic composition comprising a nicotinoyl ribosideor derivative compound that increases the level of intracellular NAD+.Exemplary skin afflictions or skin conditions that may be treated inaccordance with the methods described herein include disorders ordiseases associated with or caused by inflammation, sun damage ornatural aging. For example, the compositions find utility in theprevention or treatment of contact dermatitis (including irritantcontact dermatitis and allergic contact dermatitis), atopic dermatitis(also known as allergic eczema), actinic keratosis, keratinizationdisorders (including eczema), epidermolysis bullosa diseases (includingpenfigus), exfoliative dermatitis, seborrheic dermatitis, erythemas(including erythema multiforme and erythema nodosum), damage caused bythe sun or other light sources, discoid lupus erythematosus,dermatomyositis, psoriasis, skin cancer and the effects of naturalaging. In another embodiment, a nicotinoyl riboside or derivativecompound that increases the level of intracellular NAD+ may be used forthe treatment of wounds and/or burns to promote healing, including, forexample, first-, second- or third-degree burns and/or thermal, chemicalor electrical burns. The formulations may be administered topically, tothe skin or mucosal tissue, as an ointment, lotion, cream,microemulsion, gel, solution or the like, as further described herein,within the context of a dosing regimen effective to bring about thedesired result.

Topical formulations comprising one or more a nicotinoyl riboside orderivative compound that increases the level of intracellular NAD+ mayalso be used as preventive, e.g., chemopreventive, compositions. Whenused in a chemopreventive method, susceptible skin is treated prior toany visible condition in a particular individual.

In another embodiment, a nicotinoyl riboside or derivative compound thatincreases the level of intracellular NAD+ may be used for treating orpreventing a disease or condition induced or exacerbated by cellularsenescence in a subject; methods for decreasing the rate of senescenceof a subject, e.g., after onset of senescence; methods for extending thelifespan of a subject; methods for treating or preventing a disease orcondition relating to lifespan; methods for treating or preventing adisease or condition relating to the proliferative capacity of cells;and methods for treating or preventing a disease or condition resultingfrom cell damage or death. In certain embodiments, the method does notact by decreasing the rate of occurrence of diseases that shorten thelifespan of a subject. In certain embodiments, a method does not act byreducing the lethality caused by a disease, such as cancer.

In yet another embodiment, a nicotinoyl riboside or derivative compoundthat increases the level of intracellular NAD+ may be administered to asubject in order to generally increase the lifespan of its cells and toprotect its cells against stress and/or against apoptosis. It isbelieved that treating a subject with a compound described herein issimilar to subjecting the subject to hormesis, i.e., mild stress that isbeneficial to organisms and may extend their lifespan.

A nicotinoyl riboside or derivative compound that increases the level ofintracellular NAD+ can also be administered to subjects for treatment ofdiseases, e.g., chronic diseases, associated with cell death, in orderto protect the cells from cell death. Exemplary diseases include thoseassociated with neural cell death, neuronal dysfunction, or muscularcell death or dysfunction, such as Parkinson's disease, Alzheimer'sdisease, multiple sclerosis, amyotropic lateral sclerosis, and musculardystrophy; AIDS; fulminant hepatitis; diseases linked to degeneration ofthe brain, such as Creutzfeld-Jakob disease, retinitis pigmentosa andcerebellar degeneration; myelodysplasis such as aplastic anemia;ischemic diseases such as myocardial infarction and stroke; hepaticdiseases such as alcoholic hepatitis, hepatitis B and hepatitis C;joint-diseases such as osteoarthritis; atherosclerosis; alopecia; damageto the skin due to UV light; lichen planus; atrophy of the skin;cataract; and graft rejections. Cell death can also be caused bysurgery, drug therapy, chemical exposure or radiation exposure.

A nicotinoyl riboside or derivative compound that increases the level ofintracellular NAD+ can also be administered to a subject suffering froman acute disease, e.g., damage to an organ or tissue, e.g., a subjectsuffering from stroke or myocardial infarction or a subject sufferingfrom a spinal cord injury. A nicotinoyl riboside or derivative compoundthat increases the level of intracellular NAD+ may also be used torepair an alcoholic's liver.

Cardiovascular Disease

In another embodiment, the invention provides a method for treatingand/or preventing a cardiovascular disease by administering to a subjectin need thereof a nicotinoyl riboside or derivative compound thatincreases the level of intracellular NAD+.

Cardiovascular diseases that can be treated or prevented a nicotinoylriboside or derivative compound that increases the level ofintracellular NAD+ include cardiomyopathy or myocarditis; such asidiopathic cardiomyopathy, metabolic cardiomyopathy, alcoholiccardiomyopathy, drug-induced cardiomyopathy, ischemic cardiomyopathy,and hypertensive cardiomyopathy. Also treatable or preventable usingcompounds and methods described herein are atheromatous disorders of themajor blood vessels (macrovascular disease) such as the aorta, thecoronary arteries, the carotid arteries, the cerebrovascular arteries,the renal arteries, the iliac arteries, the femoral arteries, and thepopliteal arteries. Other vascular diseases that can be treated orprevented include those related to platelet aggregation, the retinalarterioles, the glomerular arterioles, the vasa nervorum, cardiacarterioles, and associated capillary beds of the eye, the kidney, theheart, and the central and peripheral nervous systems. The a nicotinoylriboside or derivative compound that increases the level ofintracellular NAD+ may also be used for increasing HDL levels in plasmaof an individual.

Yet other disorders that may be treated with a nicotinoyl riboside orderivative compound that increases the level of intracellular NAD+include restenosis, e.g., following coronary intervention, and disordersrelating to an abnormal level of high density and low densitycholesterol.

In one embodiment, a nicotinoyl riboside or derivative compound thatincreases the level of intracellular NAD+ may be administered as part ofa combination therapeutic with another cardiovascular agent including,for example, an anti-arrhythmic agent, an antihypertensive agent, acalcium channel blocker, a cardioplegic solution, a cardiotonic agent, afibrinolytic agent, a sclerosing solution, a vasoconstrictor agent, avasodilator agent, a nitric oxide donor, a potassium channel blocker, asodium channel blocker, statins, or a naturiuretic agent.

In one embodiment, a nicotinoyl riboside or derivative compound thatincreases the level of intracellular NAD+ may be administered as part ofa combination therapeutic with an anti-arrhythmia agent. Anti-arrhythmiaagents are often organized into four main groups according to theirmechanism of action: type I, sodium channel blockade; type II,beta-adrenergic blockade; type III, repolarization prolongation; andtype IV, calcium channel blockade. Type I anti-arrhythmic agents includelidocaine, moricizine, mexiletine, tocainide, procainamide, encainide,flecanide, tocainide, phenytoin, propafenone, quinidine, disopyramide,and flecainide. Type II anti-arrhythmic agents include propranolol andesmolol. Type III includes agents that act by prolonging the duration ofthe action potential, such as amiodarone, artilide, bretylium,clofilium, isobutilide, sotalol, azimilide, dofetilide, dronedarone,ersentilide, ibutilide, tedisamil, and trecetilide. Type IVanti-arrhythmic agents include verapamil, diltaizem, digitalis,adenosine, nickel chloride, and magnesium ions.

In another embodiment, a nicotinoyl riboside or derivative compound thatincreases the level of intracellular NAD+ may be administered as part ofa combination therapeutic with another cardiovascular agent. Examples ofcardiovascular agents include vasodilators, for example, hydralazine;angiotensin converting enzyme inhibitors, for example, captopril;anti-anginal agents, for example, isosorbide nitrate, glyceryltrinitrate and pentaerythritol tetranitrate; anti-arrhythmic agents, forexample, quinidine, procainaltide and lignocaine; cardioglycosides, forexample, digoxin and digitoxin; calcium antagonists, for example,verapamil and nifedipine; diuretics, such as thiazides and relatedcompounds, for example, bendrofluazide, chlorothiazide, chlorothalidone,hydrochlorothiazide and other diuretics, for example, fursemide andtriamterene, and sedatives, for example, nitrazepam, flurazepam anddiazepam. Other exemplary cardiovascular agents include, for example, acyclooxygenase inhibitor such as aspirin or indomethacin, a plateletaggregation inhibitor such as clopidogrel, ticlopidene or aspirin,fibrinogen antagonists or a diuretic such as chlorothiazide,hydrochlorothiazide, flumethiazide, hydroflumethiazide,bendroflumethiazide, methylchlorthiazide, trichloromethiazide,polythiazide or benzthiazide as well as ethacrynic acid tricrynafen,chlorthalidone, furosemide, musolimine, bumetanide, triamterene,amiloride and spironolactone and salts of such compounds, angiotensinconverting enzyme inhibitors such as captopril, zofenopril, fosinopril,enalapril, ceranopril, cilazopril, delapril, pentopril, quinapril,ramipril, lisinopril, and salts of such compounds, angiotensin IIantagonists such as losartan, irbesartan or valsartan, thrombolyticagents such as tissue plasminogen activator (tPA), recombinant tPA,streptokinase, urokinase, prourokinase, and anisoylated plasminogenstreptokinase activator complex (APSAC, Eminase, Beecham Laboratories),or animal salivary gland plasminogen activators, calcium channelblocking agents such as verapamil, nifedipine or diltiazem, thromboxanereceptor antagonists such as ifetroban, prostacyclin mimetics, orphosphodiesterase inhibitors. Such combination products if formulated asa fixed dose employ the compounds of this invention within the doserange described above and the other pharmaceutically active agent withinits approved dose range.

Yet other exemplary cardiovascular agents include, for example,vasodilators, e.g., bencyclane, cinnarizine, citicoline, cyclandelate,cyclonicate, ebumamonine, phenoxezyl, flunarizine, ibudilast,ifenprodil, lomerizine, naphlole, nikamate, nosergoline, nimodipine,papaverine, pentifylline, nofedoline, vincamin, vinpocetine, vichizyl,pentoxifylline, prostacyclin derivatives (such as prostaglandin E1 andprostaglandin I2), an endothelin receptor blocking drug (such asbosentan), diltiazem, nicorandil, and nitroglycerin. Examples of thecerebral protecting drug include radical scavengers (such as edaravone,vitamin E, and vitamin C), glutamate antagonists, AMPA antagonists,kainate antagonists, NMDA antagonists, GABA agonists, growth factors,opioid antagonists, phosphatidylcholine precursors, serotonin agonists,Na.sup.+/Ca.sup.2+ channel inhibitory drugs, and K.sup.+ channel openingdrugs. Examples of the brain metabolic stimulants include amantadine,tiapride, and gamma-aminobutyric acid. Examples of the anticoagulantinclude heparins (such as heparin sodium, heparin potassium, dalteparinsodium, dalteparin calcium, heparin calcium, parnaparin sodium,reviparin sodium, and danaparoid sodium), warfarin, enoxaparin,argatroban, batroxobin, and sodium citrate. Examples of the antiplateletdrug include ticlopidine hydrochloride, dipyridamole, cilostazol, ethylicosapentate, sarpogrelate hydrochloride, dilazep hydrochloride,trapidil, a nonsteroidal antiinflammatory agent (such as aspirin),beraprostsodium, iloprost, and indobufene. Examples of the thrombolyticdrug include urokinase, tissue-type plasminogen activators (such asalteplase, tisokinase, nateplase, pamiteplase, monteplase, andrateplase), and nasaruplase. Examples of the antihypertensive druginclude angiotensin converting enzyme inhibitors (such as captopril,alacepril, lisinopril, imidapril, quinapril, temocapril, delapril,benazepril, cilazapril, trandolapril, enalapril, ceronapril, fosinopril,imadapril, mobertpril, perindopril, ramipril, spirapril, andrandolapril), angiotensin II antagonists (such as losartan, candesartan,valsartan, eprosartan, and irbesartan), calcium channel blocking drugs(such as aranidipine, efonidipine, nicardipine, bamidipine, benidipine,manidipine, cilnidipine, nisoldipine, nitrendipine, nifedipine,nilvadipine, felodipine, amlodipine, diltiazem, bepridil, clentiazem,phendilin, galopamil, mibefradil, prenylamine, semotiadil, terodiline,verapamil, cilnidipine, elgodipine, isradipine, lacidipine,lercanidipine, nimodipine, cinnarizine, flunarizine, lidoflazine,lomerizine, bencyclane, etafenone, and perhexiline), .beta.-adrenalinereceptor blocking drugs (propranolol, pindolol, indenolol, carteolol,bunitrolol, atenolol, acebutolol, metoprolol, timolol, nipradilol,penbutolol, nadolol, tilisolol, carvedilol, bisoprolol, betaxolol,celiprolol, bopindolol, bevantolol, labetalol, alprenolol, amosulalol,arotinolol, befunolol, bucumolol, bufetolol, buferalol, buprandolol,butylidine, butofilolol, carazolol, cetamolol, cloranolol, dilevalol,epanolol, levobunolol, mepindolol, metipranolol, moprolol, nadoxolol,nevibolol, oxprenolol, practol, pronetalol, sotalol, sufinalol,talindolol, tertalol, toliprolol, xybenolol, and esmolol),.alpha.-receptor blocking drugs (such as amosulalol, prazosin,terazosin, doxazosin, bunazosin, urapidil, phentolamine, arotinolol,dapiprazole, fenspiride, indoramin, labetalol, naftopidil, nicergoline,tamsulosin, tolazoline, trimazosin, and yohimbine), sympathetic nerveinhibitors (such as clonidine, guanfacine, guanabenz, methyldopa, andreserpine), hydralazine, todralazine, budralazine, and cadralazine.Examples of the antianginal drug include nitrate drugs (such as amylnitrite, nitroglycerin, and isosorbide), .beta.-adrenaline receptorblocking drugs (such as propranolol, pindolol, indenolol, carteolol,bunitrolol, atenolol, acebutolol, metoprolol, timolol, nipradilol,penbutolol, nadolol, tilisolol, carvedilol, bisoprolol, betaxolol,celiprolol, bopindolol, bevantolol, labetalol, alprenolol, amosulalol,arotinolol, befunolol, bucumolol, bufetolol, buferalol, buprandolol,butylidine, butofilolol, carazolol, cetamolol, cloranolol, dilevalol,epanolol, levobunolol, mepindolol, metipranolol, moprolol, nadoxolol,nevibolol, oxprenolol, practol, pronetalol, sotalol, sufinalol,talindolol, tertalol, toliprolol, andxybenolol), calcium channelblocking drugs (such as aranidipine, efonidipine, nicardipine,bamidipine, benidipine, manidipine, cilnidipine, nisoldipine,nitrendipine, nifedipine, nilvadipine, felodipine, amlodipine,diltiazem, bepridil, clentiazem, phendiline, galopamil, mibefradil,prenylamine, semotiadil, terodiline, verapamil, cilnidipine, elgodipine,isradipine, lacidipine, lercanidipine, nimodipine, cinnarizine,flunarizine, lidoflazine, lomerizine, bencyclane, etafenone, andperhexiline) trimetazidine, dipyridamole, etafenone, dilazep, trapidil,nicorandil, enoxaparin, and aspirin. Examples of the diuretic includethiazide diuretics (such as hydrochlorothiazide, methyclothiazide,trichlormethiazide, benzylhydrochlorothiazide, and penflutizide), loopdiuretics (such as furosemide, etacrynic acid, bumetanide, piretanide,azosemide, and torasemide), K⁺ sparing diuretics (spironolactone,triamterene, and potassium can renoate), osmotic diuretics (such asisosorbide, D-mannitol, and glycerin), nonthiazide diuretics (such asmeticrane, tripamide, chlorthalidone, and mefruside), and acetazolamide.Examples of the cardiotonic include digitalis formulations (such asdigitoxin, digoxin, methyldigoxin, deslanoside, vesnarinone, lanatosideC, and proscillaridin), xanthine formulations (such as aminophylline,choline theophylline, diprophylline, and proxyphylline), catecholamineformulations (such as dopamine, dobutamine, and docarpamine), PDE IIIinhibitors (such as amrinone, olprinone, and milrinone), denopamine,ubidecarenone, pimobendan, levosimendan, aminoethylsulfonic acid,vesnarinone, carperitide, and colforsin daropate. Examples of theantiarrhythmic drug include ajmaline, pirmenol, procainamide,cibenzoline, disopyramide, quinidine, aprindine, mexiletine, lidocaine,phenyloin, pilsicainide, propafenone, flecainide, atenolol, acebutolol,sotalol, propranolol, metoprolol, pindolol, amiodarone, nifekalant,diltiazem, bepridil, and verapamil. Examples of the antihyperlipidemicdrug include atorvastatin, simvastatin, pravastatin sodium, fluvastatinsodium, clinofibrate, clofibrate, simfibrate, fenofibrate, bezafibrate,colestimide, and colestyramine. Examples of the immunosuppressantinclude azathioprine, mizoribine, cyclosporine, tacrolimus, gusperimus,and methotrexate.

Cell Death/Cancer

Tiazofurin and benzamide riboside, analogs of NAR, have been usedclinically as anticancer agents, since in vivo they could be metabolizedto form NAD+ analogs, which inhibit IMP dehydrogenase, the rate-limitingenzyme for guanine nucleotide biosynthesis. Treatment of cultured cellshas shown that tiazofurin reduces cancer cells by induction ofapoptosis.

A nicotinoyl riboside or derivative compound that increases the level ofintracellular NAD+ may be administered to subjects who have recentlyreceived or are likely to receive a dose of radiation or toxin. In oneembodiment, the dose of radiation or toxin is received as part of awork-related or medical procedure, e.g., working in a nuclear powerplant, flying an airplane, an X-ray, CAT scan, or the administration ofa radioactive dye for medical imaging; in such an embodiment, thecompound is administered as a prophylactic measure. In anotherembodiment, the radiation or toxin exposure is received unintentionally,e.g., as a result of an industrial accident, habitation in a location ofnatural radiation, terrorist act, or act of war involving radioactive ortoxic material. In such a case, the compound is preferably administeredas soon as possible after the exposure to inhibit apoptosis and thesubsequent development of acute radiation syndrome.

A nicotinoyl riboside or derivative compound may also be used fortreating and/or preventing cancer. In certain embodiments, a nicotinoylriboside or derivative compound that increases the level ofintracellular NAD+ may be used for treating and/or preventing cancer.Calorie restriction has been linked to a reduction in the incidence ofage-related disorders including cancer

A nicotinoyl riboside or derivative compound may also be used to reducethe adverse effects of drugs that are used to treat and/or preventcancer. In certain embodiments, a nicotinoyl riboside or derivativecompound that increases the level of intracellular NAD+ may be used toreduce the adverse effects of drugs that are used to treat and/orprevent cancer. Calorie restriction has been linked to a reduction inthe incidence of age-related disorders including cancer

In other embodiments, a nicotinoyl riboside or derivative compound thatdecrease the level of intracellular NAD+ may be used for treating orpreventing cancer. For example, inhibitory compounds may be used tostimulate acetylation of substrates such as p53 and thereby increaseapoptosis, as well as to reduce the lifespan of cells and organisms,render them more sensitive to stress, and/or increase theradiosensitivity and/or chemosensitivity of a cell or organism. Thus,inhibitory compounds may be used, e.g., for treating cancer. Exemplaryare those of the brain and kidney; hormone-dependent cancers includingbreast, prostate, testicular, and ovarian cancers; lymphomas, andleukemias. In cancers associated with solid tumors, a modulatingcompound may be administered directly into the tumor. Cancer of bloodcells, e.g., leukemia, can be treated by administering a modulatingcompound into the blood stream or into the bone marrow. Benign cellgrowth can also be treated, e.g., warts. Other diseases that can betreated include autoimmune diseases, e.g., systemic lupus erythematosus,scleroderma, and arthritis, in which autoimmune cells should be removed.Viral infections such as herpes, HIV, adenovirus, and HTLV-1 associatedmalignant and benign disorders can also be treated. Alternatively, cellscan be obtained from a subject, treated ex vivo to remove certainundesirable cells, e.g., cancer cells, and administered back to the sameor a different subject.

Chemotherapeutic agents that may be coadministered with modulatingcompounds described herein as having anti-cancer activity (e.g.,compounds that induce apoptosis, compounds that reduce lifespan orcompounds that render cells sensitive to stress) include:aminoglutethimide, amsacrine, anastrozole, asparaginase, bcg,bicalutamide, bleomycin, buserelin, busulfan, campothecin, capecitabine,carboplatin, carmustine, chlorambucil, cisplatin, cladribine,clodronate, colchicine, cyclophosphamide, cyproterone, cytarabine,dacarbazine, dactinomycin, daunorubicin, dienestrol, diethylstilbestrol,docetaxel, doxorubicin, epirubicin, estradiol, estramustine, etoposide,exemestane, filgrastim, fludarabine, fludrocortisone, fluorouracil,fluoxymesterone, flutamide, gemcitabine, genistein, goserelin,hydroxyurea, idarubicin, ifosfamide, imatinib, interferon, irinotecan,ironotecan, letrozole, leucovorin, leuprolide, levamisole, lomustine,mechlorethamine, medroxyprogesterone, megestrol, melphalan,mercaptopurine, mesna, methotrexate, mitomycin, mitotane, mitoxantrone,nilutamide, nocodazole, octreotide, oxaliplatin, paclitaxel,pamidronate, pentostatin, plicamycin, porfimer, procarbazine,raltitrexed, rituximab, streptozocin, suramin, tamoxifen, temozolomide,teniposide, testosterone, thioguanine, thiotepa, titanocene dichloride,topotecan, trastuzumab, tretinoin, vinblastine, vincristine, vindesine,and vinorelbine. These chemotherapeutic agents may be categorized bytheir mechanism of action into, for example, following groups:anti-metabolites/anti-cancer agents, such as pyrimidine analogs(5-fluorouracil, floxuridine, capecitabine, gemcitabine and cytarabine)and purine analogs, folate antagonists and related inhibitors(mercaptopurine, thioguanine, pentostatin and 2-chlorodeoxyadenosine(cladribine)); antiproliferative/antimitotic agents including naturalproducts such as vinca alkaloids (vinblastine, vincristine, andvinorelbine), microtubule disruptors such as taxane (paclitaxel,docetaxel), vincristin, vinblastin, nocodazole, epothilones andnavelbine, epidipodophyllotoxins (teniposide), DNA damaging agents(actinomycin, amsacrine, anthracyclines, bleomycin, busulfan,camptothecin, carboplatin, chlorambucil, cisplatin, cyclophosphamide,cytoxan, dactinomycin, daunorubicin, docetaxel, doxorubicin, epirubicin,hexamethylmelamineoxaliplatin, iphosphamide, melphalan,merchlorethamine, mitomycin, mitoxantrone, nitrosourea, paclitaxel,plicamycin, procarbazine, teniposide, triethylenethiophosphoramide andetoposide (VP16)); antibiotics such as dactinomycin (actinomycin D),daunorubicin, doxorubicin (adriamycin), idarubicin, anthracyclines,mitoxantrone, bleomycins, plicamycin (mithramycin) and mitomycin;enzymes (L-asparaginase which systemically metabolizes L-asparagine anddeprives cells which do not have the capacity to synthesize their ownasparagine); antiplatelet agents; antiproliferative/antimitoticalkylating agents such as nitrogen mustards (mechlorethamine,cyclophosphamide and analogs, melphalan, chlorambucil), ethyleniminesand methylmelamines (hexamethylmelamine and thiotepa), alkylsulfonates-busulfan, nitrosoureas (carmustine (BCNU) and analogs,streptozocin), trazenes-dacarbazinine (DTIC);antiproliferative/antimitotic antimetabolites such as folic acid analogs(methotrexate); platinum coordination complexes (cisplatin,carboplatin), procarbazine, hydroxyurea, mitotane, aminoglutethimide;hormones, hormone analogs (estrogen, tamoxifen, goserelin, bicalutamide,nilutamide) and aromatase inhibitors (letrozole, anastrozole);anticoagulants (heparin, synthetic heparin salts and other inhibitors ofthrombin); fibrinolytic agents (such as tissue plasminogen activator,streptokinase and urokinase), aspirin, COX-2 inhibitors, dipyridamole,ticlopidine, clopidogrel, abciximab; antimigratory agents; antisecretoryagents (breveldin); immunosuppressives (cyclosporine, tacrolimus(FK-506), sirolimus (rapamycin), azathioprine, mycophenolate mofetil);anti-angiogenic compounds (TNP-470, genistein) and growth factorinhibitors (vascular endothelial growth factor (VEGF) inhibitors,fibroblast growth factor (FGF) inhibitors, epidermal growth factor (EGF)inhibitors); angiotensin receptor blocker; nitric oxide donors;anti-sense oligonucleotides; antibodies (trastuzumab); cell cycleinhibitors and differentiation inducers (tretinoin); mTOR inhibitors,topoisomerase inhibitors (doxorubicin (adriamycin), amsacrine,camptothecin, daunorubicin, dactinomycin, eniposide, epirubicin,etoposide, idarubicin, irinotecan (CPT-11) and mitoxantrone, topotecan,irinotecan), corticosteroids (cortisone, dexamethasone, hydrocortisone,methylpednisolone, prednisone, and prenisolone); growth factor signaltransduction kinase inhibitors; mitochondrial dysfunction inducers andcaspase activators; chromatin disruptors.

These chemotherapeutic agents may be used by themselves with anicotinoyl riboside or derivative compound described herein as inducingcell death or reducing lifespan or increasing sensitivity to stressand/or in combination with other chemotherapeutics agents. Exemplarycombinatorial therapies for the treatment of cancer. Name Therapeuticagents ABV Doxorubicin, Bleomycin, Vinblastine ABVD Doxorubicin,Bleomycin, Vinblastine, Dacarbazine AC (Breast) Doxorubicin,Cyclophosphamide AC (Sarcoma) Doxorubicin, Cisplatin AC(Neuro-Cyclophosphamide, Doxorubicin blastoma) ACE Cyclophosphamide,Doxorubicin, Etoposide ACe Cyclophosphamide, Doxorubicin AD Doxorubicin,Dacarbazine AP Doxorubicin, Cisplatin ARAC-DNR Cytarabine, DaunorubicinB-CAVe Bleomycin, Lomustine, Doxorubicin, Vinblastine BCVPP Carmustine,Cyclophosphamide, Vinblastine, Procarbazine, Prednisone BEACOPPBleomycin, Etoposide, Doxorubicin, Cyclophosphamide, Vincristine,Procarbazine, Prednisone, Filgrastim BEP Bleomycin, Etoposide, CisplatinBIP Bleomycin, Cisplatin, Ifosfamide, Mesna BOMP Bleomycin, Vincristine,Cisplatin, Mitomycin CA Cytarabine, Asparaginase CABO Cisplatin,Methotrexate, Bleomycin, Vincristine CAF Cyclophosphamide, Doxorubicin,Fluorouracil CAL-G Cyclophosphamide, Daunorubicin, Vincristine,Prednisone, Asparaginase CAMP Cyclophosphamide, Doxorubicin,Methotrexate, Procarbazine CAP Cyclophosphamide, Doxorubicin, CisplatinCaT Carboplatin, Paclitaxel CAV Cyclophosphamide, Doxorubicin,Vincristine CAVE ADD CAV and Etoposide CA-VP16 Cyclophosphamide,Doxorubicin, Etoposide CC Cyclophosphamide, Carboplatin CDDP/VP-16Cisplatin, Etoposide CEF Cyclophosphamide, Epirubicin, FluorouracilCEPP(B) Cyclophosphamide, Etoposide, Prednisone, with orwithout/Bleomycin CEV Cyclophosphamide, Etoposide, Vincristine CFCisplatin, Fluorouracil or Carboplatin Fluorouracil CHAPCyclophosphamide or Cyclophosphamide, Altretamine, Doxorubicin,Cisplatin ChlVPP Chlorambucil, Vinblastine, Procarbazine, PrednisoneCHOP Cyclophosphamide, Doxorubicin, Vincristine, Prednisone CHOP-BLEOAdd Bleomycin to CHOP CISCA Cyclophosphamide, Doxorubicin, CisplatinCLD-BOMP Bleomycin, Cisplatin, Vincristine, Mitomycin CMF Methotrexate,Fluorouracil, Cyclophosphamide CMFP Cyclophosphamide, Methotrexate,Fluorouracil, Prednisone CMFVP Cyclophosphamide, Methotrexate,Fluorouracil, Vincristine, Prednisone CMV Cisplatin, Methotrexate,Vinblastine CNF Cyclophosphamide, Mitoxantrone, Fluorouracil CNOPCyclophosphamide, Mitoxantrone, Vincristine, Prednisone COB Cisplatin,Vincristine, Bleomycin CODE Cisplatin, Vincristine, Doxorubicin,Etoposide COMLA Cyclophosphamide, Vincristine, Methotrexate, Leucovorin,Cytarabine COMP Cyclophosphamide, Vincristine, Methotrexate, PrednisoneCooper Cyclophosphamide, Methotrexate, Fluorouracil, RegimenVincristine, Prednisone COP Cyclophosphamide, Vincristine, PrednisoneCOPE Cyclophosphamide, Vincristine, Cisplatin, Etoposide COPPCyclophosphamide, Vincristine, Procarbazine, Prednisone CP (ChronicChlorambucil, Prednisone lymphocytic leukemia) CP (OvarianCyclophosphamide, Cisplatin Cancer) CT Cisplatin, Paclitaxel CVDCisplatin, Vinblastine, Dacarbazine CVI Carboplatin, Etoposide,Ifosfamide, Mesna CVP Cyclophosphamide, Vincristine, Prednisome CVPPLomustine, Procarbazine, Prednisone CYVADIC Cyclophosphamide,Vincristine, Doxorubicin, Dacarbazine DA Daunorubicin, Cytarabine DATDaunorubicin, Cytarabine, Thioguanine DAV Daunorubicin, Cytarabine,Etoposide DCT Daunorubicin, Cytarabine, Thioguanine DHAP Cisplatin,Cytarabine, Dexamethasone DI Doxorubicin, Ifosfamide DTIC/Dacarbazine,Tamoxifen Tamoxifen DVP Daunorubicin, Vincristine, Prednisone EAPEtoposide, Doxorubicin, Cisplatin EC Etoposide, Carboplatin EFPEtoposie, Fluorouracil, Cisplatin ELF Etoposide, Leucovorin,Fluorouracil EMA 86 Mitoxantrone, Etoposide, Cytarabine EP Etoposide,Cisplatin EVA Etoposide, Vinblastine FAC Fluorouracil, Doxorubicin,Cyclophosphamide FAM Fluorouracil, Doxorubicin, Mitomycin FAMTXMethotrexate, Leucovorin, Doxorubicin FAP Fluorouracil, Doxorubicin,Cisplatin F-CL Fluorouracil, Leucovorin FEC Fluorouracil,Cyclophosphamide, Epirubicin FED Fluorouracil, Etoposide, Cisplatin FLFlutamide, Leuprolide FZ Flutamide, Goserelin acetate implant HDMTXMethotrexate, Leucovorin Hexa-CAF Altretamine, Cyclophosphamide,Methotrexate, Fluorouracil ICE-T Ifosfamide, Carboplatin, Etoposide,Paclitaxel, Mesna IDMTX/6-MP Methotrexate, Mercaptopurine, Leucovorin IEIfosfamide, Etoposie, Mesna IfoVP Ifosfamide, Etoposide, Mesna IPAIfosfamide, Cisplatin, Doxorubicin M-2 Vincristine, Carmustine,Cyclophosphamide, Prednisone, Melphalan MAC-III Methotrexate,Leucovorin, Dactinomycin, Cyclophosphamide MACC Methotrexate,Doxorubicin, Cyclophosphamide, Lomustine MACOP-B Methotrexate,Leucovorin, Doxorubicin, Cyclophosphamide, Vincristine, Bleomycin,Prednisone MAID Mesna, Doxorubicin, Ifosfamide, Dacarbazine m-BACODBleomycin, Doxorubicin, Cyclophosphamide, Vincristine, Dexamethasone,Methotrexate, Leucovorin MBC Methotrexate, Bleomycin, Cisplatin MCMitoxantrone, Cytarabine MF Methotrexate, Fluorouracil, Leucovorin MICEIfosfamide, Carboplatin, Etoposide, Mesna MINE Mesna, Ifosfamide,Mitoxantrone, Etoposide mini-BEAM Carmustine, Etoposide, Cytarabine,Melphalan MOBP Bleomycin, Vincristine, Cisplatin, Mitomycin MOPMechlorethamine, Vincristine, Procarbazine MOPP Mechlorethamine,Vincristine, Procarbazine, Prednisone MOPP/ABV Mechlorethamine,Vincristine, Procarbazine, Prednisone, Doxorubicin, Bleomycin,Vinblastine MP Melphalan, Prednisone (multiple myeloma) MP (prostateMitoxantrone, Prednisone cancer) MTX/6-MO Methotrexate, MercaptopurineMTX/6-MP/VP Methotrexate, Mercaptopurine, Vincristine, PrednisoneMTX-CDDPAdr Methotrexate, Leucovorin, Cisplatin, Doxorubicin MV (breastMitomycin, Vinblastine cancer) MV (acute Mitoxantrone, Etoposidemyelocytic leukemia) M-VAC Vinblastine, Doxorubicin, CisplatinMethotrexate MVP Vinblastine, Cisplatin Mitomycin MVPP Mechlorethamine,Vinblastine, Procarbazine, Prednisone NFL Mitoxantrone, Fluorouracil,Leucovorin NOVP Mitoxantrone, Vinblastine, Vincristine OPA Vincristine,Prednisone, Doxorubicin OPPA Add Procarbazine to OPA. PAC Cisplatin,Doxorubicin PAC-I Cisplatin, Doxorubicin, Cyclophosphamide PA-CICisplatin, Doxorubicin PC Paclitaxel, Carboplatin or Paclitaxel,Cisplatin PCV Lomustine, Procarbazine, Vincristine PE Paclitaxel,Estramustine PFL Cisplatin, Fluorouracil, Leucovorin POC Prednisone,Vincristine, Lomustine ProMACE Prednisone, Methotrexate, Leucovorin,Doxorubicin, Cyclophosphamide, Etoposide ProMACE/Prednisone,Doxorubicin, Cyclophosphamide, cytaBOM Etoposide, Cytarabine, Bleomycin,Vincristine, Methotrexate, Leucovorin, Cotrimoxazole PRoMACE/Prednisone,Doxorubicin, Cyclophosphamide, MOPP Etoposide, Mechlorethamine,Vincristine, Procarbazine, Methotrexate, Leucovorin Pt/VM Cisplatin,Teniposide PVA Prednisone, Vincristine, Asparaginase PVB Cisplatin,Vinblastine, Bleomycin PVDA Prednisone, Vincristine, Daunorubicin,Asparaginase SMF Streptozocin, Mitomycin, Fluorouracil TADMechlorethamine, Doxorubicin, Vinblastine, Vincristine, Bleomycin,Etoposide, Prednisone TCF Paclitaxel, Cisplatin, Fluorouracil TIPPaclitaxel, Ifosfamide, Mesna, Cisplatin TTT Methotrexate, Cytarabine,Hydrocortisone Topo/CTX Cyclophosphamide, Topotecan, Mesna VAB-6Cyclophosphamide, Dactinomycin, Vinblastine, Cisplatin, Bleomycin VACVincristine, Dactinomycin, Cyclophosphamide VACAdr Vincristine,Cyclophosphamide, Doxorubicin, Dactinomycin, Vincristine VADVincristine, Doxorubicin, Dexamethasone VATH Vinblastine, Doxorubicin,Thiotepa, Flouxymesterone VBAP Vincristine, Carmustine, Doxorubicin,Prednisone VBCMP Vincristine, Carmustine, Melphalan, Cyclophosphamide,Prednisone VC Vinorelbine, Cisplatin VCAP Vincristine, Cyclophosphamide,Doxorubicin, Prednisone VD Vinorelbine, Doxorubicin VelP Vinblastine,Cisplatin, Ifosfamide, Mesna VIP Etoposide, Cisplatin, Ifosfamide, MesnaVM Mitomycin, Vinblastine VMCP Vincristine, Melphalan, Cyclophosphamide,Prednisone VP Etoposide, Cisplatin V-TAD Etoposide, Thioguanine,Daunorubicin, Cytarabine 5+2 Cytarabine, Daunorubicin, Mitoxantrone 7+3Cytarabine with/, Daunorubicin or Idarubicin or Mitoxantrone “8 in 1”Methylprednisolone, Vincristine, Lomustine, Procarbazine, Hydroxyurea,Cisplatin, Cytarabine, Dacarbazine.

In addition to conventional chemotherapeutics, the a nicotinoyl ribosideor derivative compound described herein as capable of inducing celldeath or reducing lifespan can also be used with antisense RNA, RNAi orother polynucleotides to inhibit the expression of the cellularcomponents that contribute to unwanted cellular proliferation that aretargets of conventional chemotherapy. Such targets are, merely toillustrate, growth factors, growth factor receptors, cell cycleregulatory proteins, transcription factors, or signal transductionkinases.

Chemical Synthesis of Nicotinoyl Ribosides

Due to its biological significance, a number of chemical approaches havebeen used to prepare nicotinamide riboside (NAR). See Franchetti et al.,Bioorganic & Medicinal Chemistry Letters 14, 4655-4568 (2004) andTanimori et al., Bioorganic & Medicinal Chemistry Letters 12, 1135-1137(2002) both hereby incorporated by reference. However, there is still nosystematical way to synthesize NAR analogs. One embodiment of theinvention relates to an efficient method to stereoselectively synthesizeβ-NAR and its various derivatives in one pot with high yield. Comparedto the previous methods, this stereoselective method is more efficientand gives more possibilities to form various NAD+ analogs with a similarprocedure.

The synthesis of nicotinoyl riboside compounds was initiated with thepreparation of a preferred intermediate, ethyl1-[3,4-diacetyloxy-5-(acetyloxymethyl)oxolan-2-yl]-pyridine-3-carboxylate.Nicotinic acid riboside O-alkyl derivatives were prepared from there.Refluxing of the protected sugar,1,2,3,5-tetra-O-acetyl-β-D-ribofuranose with ethyl nicotinate (1.2equiv.) in methylene chloride under the catalysis of TMSOTf (1 equiv.)stereoselectively formed β-isomer in good yield (>90%). One of theadvantages for this method is that there is no need to go through thesilylation of nicotinamide. After evaporation of the solvent, ethyl1-[3,4-diacetyloxy-5-(acetyloxymethyl)oxolan-2-yl]-pyridine-3-carboxylatecould be used directly for the next step. The f-NAR and its derivativesare extremely water-soluble. Extraction of ethyl acetate from waterlayer could get rid of most non-polar side products and give 95% of pureproducts according to TLC. Ethyl1-[3,4-diacetyloxy-5-(acetyloxymethyl)oxolan-2-yl]-pyridine-3-carboxylatewas also treated with NaOCH₃/MeOH at −20° C. to form O-methyl nicotinateriboside. Other O-alkyl nicotinate ribosides were prepared similarly.O-Alkyl nicotinate ribosides can be hydrolyzed with esterase (fromporcine liver, sigma) at pH=7.0 to afford clean nicotinic acid riboside.Reverse HPLC was used to detect and purify products.

Thermodynamic study showed the reaction of ethyl1-[3,4-diacetyloxy-5-(acetyloxymethyl)oxolan-2-yl]-pyridine-3-carboxylateand NH₃/MeOH at −20° C. gave product methyl1-[3,4-diacetyloxy-5-(acetyloxymethyl)oxolan-2-yl]-pyridine-3-carboxylate,instead of β-NAR. However, without worked up, the reaction could gofurther to finally form β-NAR after it was restored at 4° C. This couldbe explained thermodynamically. Since amide is thermodynamically morestable than ester, at higher temperature, the more stable β-NAR would bethe primary product. However, the reaction with NaOCH₃ at lowertemperature, −20° C., kinetically favors the formation of O-methylnicotinate riboside.

The preparation of N-alkyl β-NAR derivatives starts from,2,3,5-tri-O-acetyl-β-phenyl nicotinate riboside (see FIG. 5) anactivated phenyl ester. Incubation of nicotinic chloride with phenol inTHE overnight at room temperature quantitatively produced pure phenylnicotinate, which was then treated with1,2,3,5-tetra-O-acetyl-β-D-ribofuranose in methylene chloride under thecatalysis of TMSOTf to give 2,3,5-tri-O-acetyl-β-phenyl nicotinateriboside. 2,3,5-Tri-O-acetyl-β-phenyl nicotinate riboside could betransformed further to prepare multiple N-alkyl NR derivatives withaddition of various primary or secondary amines. The ethyl esterintermediate was also considered to be used in building the N-alkyl NARderivatives since in literature ester-amide exchange has been considereda versatile and simple way to make amides. However, our thermodynamicstudy (see FIG. 6) showed that the incubation of the ethyl esterintermediate with N-methyl amine in methanol at −20° C. formed theO-methyl nicotinate riboside as the product, instead of the expectedN-methyl nicotinamide riboside. MALDI-MS of the product showed themolecular weight at 270 which is one mass more than the expected 269.The m/z peak at 138 rather than 137 was also detected which was due tothe methyl nicotinate piece coming after the C^(1′)—N¹ bond of themethyl ester was broken. All the information suggests that atransesterification occurs rather than the ester-amide exchange. In thelow temperature, both transesterification and ester-amide reactions werevery slow so that the reaction selectivity was kinetically controlled.The transesterification was more kinetically favored than theester-amide exchange because of the higher concentration of the solvent,methanol. Since amide is thermodynamically more stable than ester, weassume that the reaction selectivity at higher temperature would bethermodynamically controlled so as to form the more stable amideproduct. Therefore, the same reaction mixture was incubated at 4° C.However, after overnight, the riboside was totally decomposed, andN-methyl amine and free sugar were formed, which did not happened in thepreparation of β-NAR with using NH₃/MeOH at all. Decomposition maybe dueto the fact that N-alkyl amines are more basic/neutrophilic than freeammonia, which tend to attack the C-1′ position of the riboside andcause to the breaking of the C^(1′)—N¹ bond.

In order to avoid the decomposition, another reaction solvent,trifluoroethanol (TFE) has been tried. TFE has a pKa of 12.5, whichmakes it more acidic compared to solvent like methanol or ethanol. Thisacidity of the TFE decreases the neutrophilicity of N-alkyl amines,therefore helps stabilizing the C^(1′)—N¹ bond. The reaction of theethyl ester was then performed in trifluoroethanol under 4° C. for days.It was found the acidity of trifluoroethanol also moderates thenucleophilicity/basicity of N-methyl amine so that the ester-amideexchange reaction takes weeks to complete. The ester-amide exchange fromthe phenol ester in trifluroethanol at 4° C. could complete overnight.Interesting enough, we also found that 2′-, 3′- and 5′-O-acetyl groupson the sugar exhibit different deacetylation rates under the same basiccondition. After the treatment with N-methyl amine at 4° C. overnight,MALDI-MS has shown the molecular weight of the isolated product as 311instead of the expected 269. ¹H-NMR also agreed that the 5′-O-acetylgroup hasn't been taken off from the molecule. This selectivedeprotection method in fact brings up a convenient opportunity for aselective modification on the 5′ position of the ribose. After 4N ofNH₃/MeOH was added into the reaction mixture, the reaction was thencomplete after an overnight incubation at −20° C., and gave the expecteddeprotected product. All other N-alkyl nicotianmide riboside have alsobeen prepared with the similar procedure.

Compositions for Administration

The compositions comprising the active compounds (nicotinamide ribosidederivatives) include nutritional/dietary supplements and bulk-drugcompositions useful in the manufacture of pharmaceutical compositions(e.g., impure or non-sterile compositions) and pharmaceuticalcompositions (i.e., compositions that are suitable for administration toa subject) that can be used in the preparation of unit dosage forms.Such compositions optionally comprise a prophylactically ortherapeutically effective amount of a prophylactic and/or therapeuticagent disclosed herein or a combination of those agents and apharmaceutically acceptable carrier. Preferably, compositions of theinvention comprise a prophylactically or therapeutically effectiveamount of the active compound and another therapeutic or prophylacticagent, and a pharmaceutically acceptable carrier. These compositions maycontain between 0.1-99% of the active ingredient

In a specific embodiment, the term “pharmaceutically acceptable” meansapproved by a regulatory agency of the Federal or a state government orlisted in the U.S. Pharmacopeia or other generally recognizedpharmacopeia for use in animals, and more particularly in humans. Theterm “carrier” refers to a diluent, adjuvant, excipient, or vehicle withwhich the active compound is administered. Such pharmaceutical vehiclescan be liquids, such as water and oils, including those of petroleum,animal, vegetable or synthetic origin, such as peanut oil, soybean oil,mineral oil, sesame oil and the like. The pharmaceutical vehicles can besaline, gum acacia, gelatin, starch paste, talc, keratin, colloidalsilica, urea, and the like. In addition, auxiliary, stabilizing,thickening, lubricating and coloring agents can be used. Whenadministered to a subject, the pharmaceutically acceptable vehicles arepreferably sterile. Water can be the vehicle when the active compound isadministered intravenously. Saline solutions and aqueous dextrose andglycerol solutions can also be employed as liquid vehicles, particularlyfor injectable solutions. Suitable pharmaceutical vehicles also includeexcipients such as starch, glucose, lactose, sucrose, gelatin, malt,rice, flour, chalk, silica gel, sodium stearate, glycerol monostearate,talc, sodium chloride, dried skim milk, glycerol, propyleneglycol,water, ethanol and the like. The present compositions, if desired, canalso contain minor amounts of wetting or emulsifying agents, or pHbuffering agents.

The present compositions can take the form of solutions, suspensions,emulsion, tablets, pills, pellets, capsules, capsules containingliquids, powders, sustained-release formulations, suppositories,emulsions, aerosols, sprays, suspensions, or any other form suitable foruse. In one embodiment, the pharmaceutically acceptable vehicle is acapsule (see e.g., U.S. Pat. No. 5,698,155).

In a preferred embodiment, the active compound and optionally anothertherapeutic or prophylactic agent are formulated in accordance withroutine procedures as pharmaceutical compositions adapted forintravenous administration to human beings. Typically, the activecompound for intravenous administration are solutions in sterileisotonic aqueous buffer. Where necessary, the compositions can alsoinclude a solubilizing agent. Compositions for intravenousadministration can optionally include a local anesthetic such aslignocaine to ease pain at the site of the injection. Generally, theingredients are supplied either separately or mixed together in unitdosage form, for example, as a dry lyophilized powder or water freeconcentrate in a hermetically sealed container such as an ampoule orsachette indicating the quantity of active agent. Where the activecompound is to be administered by infusion, it can be dispensed, forexample, with an infusion bottle containing sterile pharmaceutical gradewater or saline. Where the active compound is administered by injection,an ampoule of sterile water for injection or saline can be provided sothat the ingredients can be mixed prior to administration.

Compositions for oral delivery can be in the form of tablets, lozenges,aqueous or oily suspensions, granules, powders, emulsions, capsules,syrups, or elixirs, for example. Orally administered compositions cancontain one or more optional agents, for example, sweetening agents suchas fructose, aspartame or saccharin; flavoring agents such aspeppermint, oil of wintergreen, or cherry; coloring agents; andpreserving agents, to provide a pharmaceutically palatable preparation.Moreover, where in tablet or pill form, the compositions can be coatedto delay disintegration and absorption in the gastrointestinal tractthereby providing a sustained action over an extended period of time.Selectively permeable membranes surrounding an osmotically activedriving compound are also suitable for an orally administered of theactive compound. In these later platforms, fluid from the environmentsurrounding the capsule is imbibed by the driving compound, which swellsto displace the agent or agent composition through an aperture. Thesedelivery platforms can provide an essentially zero order deliveryprofile as opposed to the spiked profiles of immediate releaseformulations. A time delay material such as glycerol monostearate orglycerol stearate can also be used. Oral compositions can includestandard vehicles such as mannitol, lactose, starch, magnesium stearate,sodium saccharine, cellulose, magnesium carbonate, and the like. Suchvehicles are preferably of pharmaceutical grade.

Further, the effect of the active compound can be delayed or prolongedby proper formulation. For example, a slowly soluble pellet of theactive compound can be prepared and incorporated in a tablet or capsule.The technique can be improved by making pellets of several differentdissolution rates and filling capsules with a mixture of the pellets.Tablets or capsules can be coated with a film that resists dissolutionfor a predictable period of time. Even the parenteral preparations canbe made long-acting, by dissolving or suspending the compound in oily oremulsified vehicles which allow it to disperse only slowly in the serum.

Compositions for use in accordance with the present invention can beformulated in conventional manner using one or more physiologicallyacceptable carriers or excipients.

Thus, the compound and optionally another therapeutic or prophylacticagent and their physiologically acceptable salts and solvates can beformulated into pharmaceutical compositions for administration byinhalation or insufflation (either through the mouth or the nose) ororal, parenteral or mucosol (such as buccal, vaginal, rectal,sublingual) administration. In one embodiment, local or systemicparenteral administration is used.

For oral administration, the compositions can take the form of, forexample, tablets or capsules prepared by conventional means withpharmaceutically acceptable excipients such as binding agents (e.g.,pregelatinised maize starch, polyvinylpyrrolidone or hydroxypropylmethylcellulose); fillers (e.g., lactose, microcrystalline cellulose orcalcium hydrogen phosphate); lubricants (e.g., magnesium stearate, talcor silica); disintegrants (e.g., potato starch or sodium starchglycolate); or wetting agents (e.g., sodium lauryl sulphate). Thetablets can be coated by methods well known in the art. Liquidpreparations for oral administration can take the form of, for example,solutions, syrups or suspensions, or they can be presented as a dryproduct for constitution with water or other suitable vehicle beforeuse. Such liquid preparations can be prepared by conventional means withpharmaceutically acceptable additives such as suspending agents (e.g.,sorbitol syrup, cellulose derivatives or hydrogenated edible fats);emulsifying agents (e.g., lecithin or acacia); non-aqueous vehicles(e.g., almond oil, oily esters, ethyl alcohol or fractionated vegetableoils); and preservatives (e.g., methyl or propyl-β-hydroxybenzoates orsorbic acid). The preparations can also contain buffer salts, flavoring,coloring and sweetening agents as appropriate.

Preparations for oral administration can be suitably formulated to givecontrolled release of the active compound.

For buccal administration the compositions can take the form of tabletsor lozenges formulated in conventional manner.

For administration by inhalation, the compositions for use according tothe present invention are conveniently delivered in the form of anaerosol spray presentation from pressurized packs or a nebulizer, withthe use of a suitable propellant, e.g., dichlorodifluoromethane,trichlorofluoromethane, dichlorotetrafluoroethane, carbon dioxide orother suitable gas. In the case of a pressurized aerosol the dosage unitcan be determined by providing a valve to deliver a metered amount.Capsules and cartridges of e.g., gelatin for use in an inhaler orinsufflator can be formulated containing a powder mix of the compoundand a suitable powder base such as lactose or starch.

The compositions can be formulated for parenteral administration byinjection, e.g., by bolus injection or continuous infusion. Formulationsfor injection can be presented in unit dosage form, e.g., in ampoules orin multi-dose containers, with an added preservative. The pharmaceuticalcompositions can take such forms as suspensions, solutions or emulsionsin oily or aqueous vehicles, and can contain formulatory agents such assuspending, stabilizing and/or dispersing agents. Alternatively, theactive ingredient can be in powder form for constitution with a suitablevehicle, e.g., sterile pyrogen-free water, before use.

The compositions can also be formulated in rectal compositions such assuppositories or retention enemas, e.g., containing conventionalsuppository bases such as cocoa butter or other glycerides.

In addition to the formulations described previously, the compositionscan also be formulated as a depot preparation. Such long actingformulations can be administered by implantation (for examplesubcutaneously or intramuscularly) or by intramuscular injection. Thus,for example, the pharmaceutical compositions can be formulated withsuitable polymeric or hydrophobic materials (for example as an emulsionin an acceptable oil) or ion exchange resins, or as sparingly solublederivatives, for example, as a sparingly soluble salt.

In other embodiments of the invention, radiation therapy agents such asradioactive isotopes can be given orally as liquids in capsules or as adrink. Radioactive isotopes can also be formulated for intravenousinjection. The skilled oncologist can determine the preferredformulation and route of administration.

The compositions can, if desired, be presented in a pack or dispenserdevice that can contain one or more unit dosage forms containing theactive ingredient. The pack can for example comprise metal or plasticfoil, such as a blister pack. The pack or dispenser device can beaccompanied by instructions for administration.

In certain preferred embodiments, the pack or dispenser contains one ormore unit dosage forms containing no more than the recommended dosageformulation as determined in the Physician's Desk Reference (56^(th) ed.2002, herein incorporated by reference in its entirety).

Methods of administering the active compound and optionally anothertherapeutic or prophylactic agent include, but are not limited to,parenteral administration (e.g., intradermal, intramuscular,intraperitoneal, intravenous and subcutaneous), epidural, and mucosal(e.g., intranasal, rectal, vaginal, sublingual, buccal or oral routes).In a specific embodiment, the active compound and optionally anotherprophylactic or therapeutic agents are administered intramuscularly,intravenously, or subcutaneously. The active compound and optionallyanother prophylactic or therapeutic agent can also be administered byinfusion or bolus injection and can be administered together with otherbiologically active agents. Administration can be local or systemic. Theactive compound and optionally the prophylactic or therapeutic agent andtheir physiologically acceptable salts and solvates can also beadministered by inhalation or insufflation (either through the mouth orthe nose). In a preferred embodiment, local or systemic parenteraladministration is used.

In specific embodiments, it can be desirable to administer the activecompound locally to the area in need of treatment. This can be achieved,for example, and not by way of limitation, by local infusion duringsurgery, topical application, e.g., in conjunction with a wound dressingafter surgery, by injection, by means of a catheter, by means of asuppository, or by means of an implant, said implant being of a porous,non-porous, or gelatinous material, including membranes, such assilastic membranes, or fibers. In one embodiment, administration can beby direct injection at the site (or former site) of an atheroscleroticplaque tissue.

Pulmonary administration can also be employed, e.g., by use of aninhaler or nebulizer, and formulation with an aerosolizing agent, or viaperfusion in a fluorocarbon or synthetic pulmonary surfactant. Incertain embodiments, the active compound can be formulated as asuppository, with traditional binders and vehicles such astriglycerides.

In another embodiment, the active compound can be delivered in avesicle, in particular a liposome.

In yet another embodiment, the active compound can be delivered in acontrolled release system. In one embodiment, a pump can be used. Inanother embodiment, polymeric materials can be used.

The amount of the active compound that is effective in the treatment orprevention of heart conditions can be determined by standard researchtechniques. For example, the dosage of the active compound which will beeffective in the treatment or prevention of heart conditions can bedetermined by administering the active compound to an animal in a modelsuch as, e.g., the animal models known to those skilled in the art. Inaddition, in vitro assays can optionally be employed to help identifyoptimal dosage ranges.

Selection of a particular effective dose can be determined (e.g., viaclinical trials) by a skilled artisan based upon the consideration ofseveral factors which will be known to one skilled in the art. Suchfactors include the disease to be treated or prevented, the symptomsinvolved, the subject's body mass, the subject's immune status and otherfactors known by the skilled artisan.

The precise dose to be employed in the formulation will also depend onthe route of administration, and the seriousness of the disease-relatedwasting, and should be decided according to the judgment of thepractitioner and each subject's circumstances. Effective doses can beextrapolated from dose-response curves derived from in vitro or animalmodel test systems.

The dose of the active compound to be administered to a subject, such asa human, is rather widely variable and can be subject to independentjudgment. It is often practical to administer the daily dose of theactive compound at various hours of the day. However, in any given case,the amount of the active compound administered will depend on suchfactors as the solubility of the active component, the formulation used,subject condition (such as weight), and/or the route of administration.

The general range of effective amounts of the active compound alone orin combination with another prophylactic or therapeutic agent(s) arefrom about 0.001 mg/day to about 1000 mg/day, more preferably from about0.001 mg/day to 750 mg/day, more preferably from about 0.001 mg/day to500 mg/day, more preferably from about 0.001 mg/day to 250 mg/day, morepreferably from about 0.001 mg/day to 100 mg/day, more preferably fromabout 0.001 mg/day to 75 mg/day, more preferably from about 0.001 mg/dayto 50 mg/day, more preferably from about 0.001 mg/day to 25 mg/day, morepreferably from about 0.001 mg/day to 10 mg/day, more preferably fromabout 0.001 mg/day to 1 mg/day. Of course, it is often practical toadminister the daily dose of compound in portions, at various hours ofthe day. However, in any given case, the amount of compound administeredwill depend on such factors as the solubility of the active component,the formulation used, subject condition (such as weight), and/or theroute of administration.

A popular cancer drug is taxol. Typical dosage ranges of taxol includeless than 10 mg to 100 mg or more. Particular doses of taxol includeabout 5 mg, 10 mg, 20 mg, 40 mg, 50 mg, 60 mg, 80 mg, 100 mg, 150 mg,200 mg. Typically, these are daily dosages. Generally, higher dosagesare less preferred because of potential gastric disturbances.Therapeutic dosages may range between 40 to 80 mg per day when tolerableby a subject.

The invention provides for any method of administrating lower doses ofknown agents (e.g., taxol) than previously thought to be useful for theprevention or treatment of cancer.

The invention provides a pharmaceutical pack or kit comprising one ormore containers containing an active compound and optionally one or moreother prophylactic or therapeutic agents useful for the prevention ortreatment of cancer. The invention also provides a pharmaceutical packor kit comprising one or more containers containing one or more of theingredients of the pharmaceutical compositions. Optionally associatedwith such container(s) can be a notice in the form prescribed by agovernmental agency regulating the manufacture, use or sale ofpharmaceuticals or biological products, which notice reflects approvalby the agency of manufacture, use or sale for human administration; orinstructions for the composition's use.

The present invention provides kits that can be used in the abovemethods. In one embodiment, a kit comprises the active compound, in oneor more containers, and optionally one or more other prophylactic ortherapeutic agents useful for the treatment of cancer, in one or morecontainers.

EXAMPLES Example 1: Synthesis of 2′,3′, 5′-Triacetyl ethyl nicotinateriboside (ethyl1-[3,4-diacetyloxy-5-(acetyloxymethyl)oxolan-2-yl]-pyridine-3-carboxylate)

One equivalent of trimethylsilyl trifluoromethanesulfonate (TMSOTf) wasslowly added into Ethyl nicotinate (0.9 mL, 6.6 moL) and1,2,3,5-tetra-O-acetyl-β-D-ribofuranose (1.4 g, 4.4 mmol) in 50 mLanhydrous methylene chloride at room temperature. The mixture was thenheated to reflux for 8 hour. TLC stained with H₂SO₄ in MeOH showed thedisappearance of 1,2,3,5-tetra-O-acetyl-β-D-ribofuranose, and almostpure product. The product could be used directly for the next step aftermethylene chloride was evaporated.

Example 2. Synthesis of β-NAR (Nicotinamide Riboside)

2′,3′, 5′-Triacetyl ethyl nicotinate riboside (180 mg, 0.44 mmol) wasadded into 4.7 mL of 4N NH₃/MeOH on ice. After mixed well, the reactionwas stored at 4° C. overnight. After methanol was removed in vacuum, theresidue was dissolved in water, and extracted with ethyl acetate threetimes to get rid of the non-polar impurity. The water layer was thenconcentrated and injected into reverse HPLC for purification (watersxTerra Prep RP18© column, 2 mL/min, 20 mM ammonium acetate).

Example 3. O-alkyl β-nicotinate ribosides

2′,3′, 5′-Triacetyl ethyl nicotinate riboside (25 mg, 0.61 μmol) wasadded into 0.9 mL of NaOMe/MeOH (255 mM) or NaOEt/EtOH (312 mM) on ice.After mixed well, the reaction was stored at −20° C. overnight. Thereaction was quenched with addition of acetic acid to pH=7. Afterorganic solvent was removed in vacuum, the residue was dissolved inwater, and extracted with cyclohexane to get rid of the non-polarimpurity. The water layer was then concentrated and injected intoreverse HPLC for purification.

TABLE 1 Entry (FIG. 1) R Temp. 14 —OCH₃ −20° C. 15 —OCH₂CH₃ −20° C. 16—OCH₂CH₂OH −20° C.

Example 4. N-Alkyl β-Nicotinamide Ribosides

Nicotinic chloride (1.78 g, 10 mmoL) was added to phenol (1.29 g, 12mmoL) in 10 mL of THF. Triethylamine (5 mL) and pyridine (5 mL) werethen added to the mixture, which was then stirred at room temperatureovernight. After THF was evaporated, the mixture was then washed withethyl acetate in water for three times. The organic layer was thenpurified with silica column (Hexane: EtOAC=4:1) to give pure product.The riboside was prepared according to the procedure provided inExample 1. This phenol ester (25 mg, 0.61 μmol) was added to variousamines (400 μM) in 0.9 mL of trifluoroethanol. After mixed well, thereaction was stored at 4° C. overnight. On the next day, 4 N of NH₃/MeOHwas added into the reaction mixture, and the reaction was stored at −20°C. overnight. After the reaction was quenched with addition of HCl tomake pH<7, the organic solvent was removed in vacuum. The residue wasdissolved in water and isolated with a Octadecyl-C18 disposableextraction.

TABLE 2 Entry Solvent Yield (Figure 1) Intermediate Temp. Product (%)

3 R′ = C₂H₅ MeOH R = NH₂ 85   4° C. 4 R′ = C₂H₅ MeOH R = OCH₃ 85 −20° C.5 R′ = C₂H₅ EtOH R = OC₂H₅ 81 −20° C. 6 R′ = C₂H₅ R = OH 80 9 R′ = C₆H₅TFE R = NHCH₃ 80  4° C. 10 R′ = C₆H₅ TFE R = NHC₂H₅ 80  4° C. 11 R′ =C₆H₅ TFE R = NHCH₂C 80  4° C. H = CH₂ 12 R′ = C₆H₅ TFE R = NHC₂H₄N 55 4° C. H₂ 13 R′ = C₆H₅ TFE R = NH(CH₃)₂ 52  4° C.

Example 5. Synthesis of β-Nicotinic acid riboside(1-[3,4-dihydroxy-5-(hydroxymethyl) oxolan-2-yl]pyridine-5-carboxylicacid)

Methyl1-[3,4-diacetyloxy-5-(acetyloxymethyl)oxolan-2-yl]-pyridine-3-carboxylatewas dissolved into phosphate buffer (150 mM, pH=7.0). 10 uL of esterasewas added to the mixture, and mixed well. The reaction was run at 25° C.overnight. HPLC injection showed 98% pure product.

Example 6. Increase of Cellular NAD+ Levels Grown on NicotinamideRiboside (NAR) Enriched Medium

Mouse embryonic stem cells were plated onto cell culture flasks (250 mL,75 cm²) and grown to 50% confluence in 48 hours after passage. At 48hours 2 flasks were set aside as controls and 2 flasks were used todetermine effect of NAR on NAD+ biosynthesis. Cells were treated with 1MHClO₄, and a specific amount of 180-NAM (nicotinamide mononucleotide)and ¹⁸O-NAD+. Samples were fractionated by HPLC and the fractionscontaining NAM and NAD+ were analyzed by ESI-MS and MALDI-MSrespectively. Peak ratio (¹⁶O/¹⁸O) was used to quantitate NAM and NAD+amounts in each sample. It was found that NAR treated cells exhibit ahigher concentration of NAD+ compared to the controls. According to ourmeasurement, the NAD+ level in NAR treated cells is 3886.6 pmoL/mg cellprotein and nucleic acid content, and the NAD+ in control cells is only1782.2 pmoL/mg cell protein and nucleic acid content. Cell protein andnucleic acid content were measured separately (see FIG. 3).

Example 7. Effect of Nicotinamide Riboside (NAR) on Mms Mediated CellDeath

Mouse embryonic stem cells are passed into 7 flasks (250 mL, 75 cm²)with 20 mL of media [For 500 mL: Dulbeccos modification of Eagles media2 mM glutamine, 10% ES qualified Fetal Bovine Serum to 10%,Penicillin/streptomycin (5 mL 100×), Non-essential amino acids (5 mL100×), Sodium pyruvate (5 mL 100×), 4 μL beta-mercapto-ethanol andleukemia inhibitory factor final conc. 1000 units/mL], named from #1 to#7. After 2 days incubation at 37° C. with 5% CO₂, NAR was added toflask #3, 5, 7 to make final concentration at 1 mM, and cells wereincubated for another day. Mms (200 mM) was added to flask #2, 3, 4, 5,6, 7 to make final concentration as 1 mM. Cell flask #1 is kept as acontrol, and used for passage. After 3 hrs incubation, fresh media waschanged for flask #4, 5, 6, 7. And Cell #2, and 3 are harvested. After 6hrs treatment of mms, Cell #4, 5 are harvested. And Cell #6, 7 areharvested at 9 hrs treatment of mms. The harvest cells (living ones) arecounted for the percentage of death with trypan blue (see FIG. 2).

Example 8. Cell Survival

Mouse embryonic stem cells are passed into 2 flasks (250 mL, 75 cm²)with 20 mL of media, named #A and #B. After 2 days incubation at 37° C.with 5% CO₂, NAR was added to flask #A to make final concentration at 1mM, and both cells were incubated for another day. Mms (200 mM) wasadded to both flask #A and #B to make final concentration as 1 mM. After3 hrs incubation, each cell culture (with/without NR treated) is passedto two new flasks with/without 1 mM of NAR. After two days incubation,count the culture survival in each of these 4 new flasks: mms/controlmedia, mms/NAR media; mms+NAR/control media; mms+NAR/NAR media. It wasfound after mms treatment, the cultured cells are seriously damaged andonly have a very low survival after passage. However, compared toothers, NAR treated cell culture provides the highest culture numbers.Cell counting by haemocytometry was used to determine survival versuscell number (see FIG. 4).

Example 9. Effects of Statins on Cell Viability and Mitigation ofToxicity by Nicotinamide Riboside

Equal numbers of mouse embryonic stem cells are passed into two 6 welltissue culture plates (10 cm² each well) with 3 mL of media [For 500 mL:Dulbeccos modification of Eagles media 2 mM glutamine, 10% ES qualifiedFetal Bovine Serum to 10%, Penicillin/streptomycin (5 mL 100×),Non-essential amino acids (5 mL 100×), Sodium pyruvate (5 mL 100×), 4 μLbeta-mercapto-ethanol and leukemia inhibitory factor final conc. 1000units/mL], named from #1 to #6. After 2 days incubation at 37° C. with5% CO₂, different concentrations of lovastatin 0, 50, 250, 800, 2000 and10000 nM was added to wells 1-6 on each plate respectively. The twoplates were then renamed NR and no NR to represent treatment and lack oftreatment with the compound nicotinamide riboside (NR). Each well in theNR plate was treated with 10 μL 150 mM NR to make a final concentrationof NR in each well of 500 μM. At 2, 4 and 7 days cells were assayed bycell counting and by MTT assay after trypinizing cells to detach themfrom plates. At 2 and 4 days cells were reseeded from each individualwell into new wells in new 6 well plates and treated immediately withthe same concentration of NR and statin as the cells from theoriginating well. The seeded cells were identical in number of viablecells in each case as determined by cell counting an appropriatedilution of the cells trypsinized from each well.

The cell viability at each time point was plotted against statinconcentration to determine the IC50 of the statin in the NR and the noNR plate where IC50 is defined as the concentration of statin thatreduces cell viability by 50% under the experimental conditions. Thestatin typically reduces cell viability by 50% at a concentrationbetween 130 nM-300 nM at each time point for cells not treated with NR(FIG. 8). In the presence of NR the IC50 of the statin increasesmoderately at 2 days treatment and two to three fold after 4 and 7 daystreatment (FIG. 8). This shows that NR mitigates statin toxicity in asignificant way with NR available in cell media.

The cell viability as measured by cell counting and MTT assay issignificantly higher in the NR treated group versus no NR at virtuallyevery statin concentration, resulting in a higher IC50 for statins inthe presence of NR (566 nM, determined by cell counting; 656 nMdetermined by MTT assay) versus the absence of NR (248 nM, determined bycell counting; 216 nM determined by MTT assay). Notably at the higheststatin concentration measured, 10 μM, significant percentages of viablecells remain in the NR treated group as measured by both cell countingand MTT assay, whereas the cell viability in the no NR group is almostcompletely non-viable. The data analysis used was analogous in the 2 and4 day results shown graphically in FIG. 8.

Example 10. Evaluation of Different Nicotinate Riboside Derivatives asBioavailable Forms of Niacin that Stimulate NAD+ Synthesis in Cells

Hela cells were seeded on a 6-well plate at 37° C. with 5% CO₂ for 24hrs. Synthesized compounds (NAR: Nicotinic acid riboside, O-ethyl-NR:ethyl-nicotinate riboside, Intermediate: TAENR,triacetyl-ethyl-nicotinate riboside, See FIG. 10) were added to thecells separately after 24 hours (no compound added for control). Thecells were then incubated for another 24 hrs before harvest. Cells weretrypsinized and an aliquot removed for counting. Cells were pelleted andthen were dissolved into 100 uL of 7% perchloric acid, and 535.5 pmoL of¹⁸O-NAD+ (95% isotopically enriched) was added to each sample. Aftermixing thoroughly, samples were pelleted and then neutralized with NaOHto make pH=7˜8. Samples were then centrifuged for 3 minutes and thesupernatant (100 uL) was injected onto a C-18 column (Hitachi HPLC L2130pump L2450 diode array detector) and separated to isolate the peakcontaining NAD+. The elution was collected during the NAD+ peak and thesolution frozen and lyophilized. The sample was assayed by MALDI-MS inpositive ion detection mode to determine the NAD+ content (Table 3) bymeasuring the m/z=664 (¹⁶O-NAD+ and 666 (¹⁸O-NAD+) peaks.

The experiment suggests that different forms of nicotinate ribosides areavailable as metabolizable sources of niacin and that these chemicalforms of niacin of novel composition have the effect of substantiallyincreasing NAD+ concentrations in cells as determined by a quantitativeMS assay. Increases in NAD+ concentrations within cells can lead toclinical benefits. The minimal increase in NAD+ concentration in cellswe observed was from 618 μmol per million cells (Table 3, and bar chartFIG. 9) in untreated cells to 1004 μmol per million cells in the case oftreatment with 500 micromolar treatment of the compound O-ethylnicotinate riboside. This represents a 62% increase over the controlNAD+ content. Increasing the concentration of this compound to 800micromolar led to an increase in NAD+ concentration of approximatelythree fold (Table 3). Similarly nicotinic acid riboside and the fullyesterified compound tri-acetyl-ethylnicotinate riboside was able toincrease NAD+ concentrations in cells by 60% and three fold respectively(Table 3). These data demonstrate that these derivatives are metabolizedby cells and increase NAD+ concentrations in cells in a substantivemanner (60-300%). These compounds increase intracellular NAD+.

TABLE 3 NAD+ content (pmoL) in ES cells after treated with differentcompounds [NAD+]/ ¹⁶O— ¹⁸O— Cell 10{circumflex over ( )}6 664/ NAD+ NAD+number cell Compounds 666 (pmol) (pmol) (*10{circumflex over ( )}6)(pmoL) NAR (669 uM) 4.11 2201.2 535.5 2.04 1079 O-ethyl NR (500 uM) 2.021084.1 535.5 1.08 1004 O-ethyl NR (803 uM) 4.90 2622.7 535.5 1.17 2242TAENR (750 uM) 8.28 4432.2 535.5 2.16 2052 Control 2.01 1075.5 535.51.74 618 NAR: Nicotinic acid riboside, O-ethyl-NR: ethyl-nicotinateriboside, TAENR, triacetyl-ethyl-nicotinate riboside

Example 11. Reduction in Statin Toxicity

ES cells were growing on four 6-well plate at 37° C. with 5% CO₂ for 24hrs. The cells were then treated with 0, 800 or 5000 nM lovastatin(final concentration) and also treated with one of 500 μM NAR, 1 mMOENR, 1 mM TAENR or no compound. After 48 hours cells media was removedand cells were harvested by trypsinizing. Cells were counted byhaemocytometry and using propidium iodide staining to assess forviability. Cells were counted for no statin treatment and 800 nM and 5μM statin treatment in each group. Cell death percentage was determinedby the viable cells in each group divided by the cell numbers in thecorresponding no statin treated group. Cell counts were similar incontrols from each group (no statin) indicating low toxicity of theadded nicotinate riboside compounds to the treated cells.

FIG. 10 shows data suggesting that statin toxicity can be mitigated bynicotinate riboside compounds. The lower cell death percentages showthat nicotinate riboside compounds prevent cells from succumbing totoxic statin treatment. We envision that these compounds can enter NAD+metabolism as demonstrated by our study of the potency of thesemolecules as NAD+ precursors and thereby protect cells from stress ortoxicity. Thus, various degenerative disorders resulting from diseaseprocesses can be treated by using these compounds.

Example 12. Toxicity of Selected Nicotinamide Riboside Derivatives

To 96 well plates, mouse ES Cells were incubated in cell culture mediafor a 24 hour period with no compound (control) dimethyl nicotinamideriboside (1 mM) or N-allyl nicotinamide riboside (1 mM). After a 24 hourperiod cells were treated with MTT reagent and further incubated at 37°C. for three additional hours. After this time, media was then removedby pipet and DMSO added to solubilize dye. Purple color was measured at590 nm in a plate reader to assess cell viability. All samples were runin duplicate and the average MTT reading ratioed against the averagereading versus control. N-allyl nicotinamide riboside is able to reducecell proliferation in comparison with controls that are not treated withcompound (FIG. 12).

Example 13. Chemical Synthesis and Characterization

All the organic solvents and reagents were purchased from Sigma-AldrichCorporation and VWR Scientific and used without further purification.The HPLC system consists of a Hitachi EZChrom Elite HPLC system with aL2450 diode array as a detector. The purification system employed amobile phase system involving 20 mM of ammonium acetate buffer with aflow rate of 2.0 mL/min and a Waters Delta PAK Cis (15 μm, 300×7.8 mm)column. MALDI-mass spectra were obtained from Proteomics Resource centerat Rockefeller University with using the PerSeptive Voyager DERPMALDI-TOF Mass Spectrometer. HRMS spectra were obtained from the HunterCollege Mass Spectrometry Facility. ¹H and ¹³C NMR spectra were recordedon Bruker NMR spectrometer operating at 400 MHz (DPX Avance),respectively. ¹H and ¹³C chemical shifts are expressed in ppm withrespect to the standard deuterium solvent peaks.

Phenyl Nicotinate:

Nicotinic chloride (1.78 g, 10 mmoL) was added to phenol (1.29 g, 12mmoL) in 10 mL of tetrahydrofuran (THF). Triethylamine (5 mL) andpyridine (5 mL) were then added to the mixture, which was then stirredat room temperature overnight. After THF was evaporated, the mixture waswashed with ethyl acetate in water for three times. The ethyl acetate(EtOAC) layer was then purified with silica column (Hexane: EtOAC=4:1)to give pure product. 1H NMR (400 MHz, CDCl₃): δ=9.40 (s, 1H), 8.86 (d,1H), 8.46 (d, 1H), 7.45 (m, 3H), 7.32 (d, 1H), 7.24 (m, 2H).

2′,3′,5′-Triacetyl Ethyl Nicotinate Riboside (2)

1 equiv. of trimethylsilyl trifluoromethanesulfonate (TMSOTf) was slowlyadded into ethyl nicotinate (0.9 mL, 6.6 moL) and1,2,3,5-tetra-O-acetyl-β-D-ribofuranose (1.4 g, 4.4 mmol) in 50 mLanhydrous methylene chloride at room temperature. The mixture was thenheated to reflux for 8 hr. TLC (MeOH:MeOH:TEA=5:0.3:0.05) stained with10% of H₂SO₄ in MeOH showed the disappearance of the ribofuranose, andan almost pure product. The product 2 was then used directly for thenext step after methylene chloride was evaporated. MS: M/Z (%): 410.06(14, [M]⁺), 259.0 (100), 138.9 (22).

2′,3′,5′-Triacetyl Phenyl Nicotinate Riboside (8)

1 equiv. of trimethylsilyl trifluoromethanesulfonate (TMSOTf) was slowlyadded into phenyl nicotinate (100 mg, 0.41 mmoL) and1,2,3,5-tetra-O-acetyl-β-D-ribofuranose (159 mg, 0.5 mmol) in 15 mLanhydrous methylene chloride at room temperature. The mixture was thenheated to reflux for 5 hr. TLC (MeOH:MeOH:TEA=5:0.3:0.05) stained with10% of H₂SO₄ in MeOH showed the disappearance of 1, and almost pureproduct. The product could be used directly for the next step aftermethylene chloride was evaporated. MS: M/Z (%): 458.1 (12, [M]⁺), 139.0(64), 200.0 (76).

β-Nicotinamide Riboside (3):

Compound 2 (180 mg, 0.44 mmol) was added into 4.7 mL of 4N of NH₃/MeOHon ice. After mixed well, the reaction was stored at 4° C. overnight.After methanol was removed in vacuum, the residue was dissolved inwater, and extracted with ethyl acetate for three times to get rid ofthe non-polar impurity. The water layer was then concentrated andinjected into reverse HPLC for purification. 1H NMR (400 MHz, D₂O):δ=9.60 (s, 1H), 9.31 (d, 1H), 8.92 (d, 1H), 8.16 (t, 1H), 6.06 (d, 1H),4.31 (m, 2H), 4.19 (t, 1H), 3.90 (dd, 1H), 3.74 (dd, 1H). 13C NMR (300MHz, D₂O): δ=146.0, 143.0, 141.3, 128.7, 100.2, 88.1, 77.7, 70.2, 60.6.MS: 255 (67, [M]⁺), 133.0 (4), 123.0 (82). HRMS (ESI) m/z [M]⁺ calcd forC₁₁H₁₅N₂O₅ 255.09755, found 255.09801.

O-Alkyl β-Nicotinic Riboside (4, 5):

Compound 2 (25 mg, 0.61 μmol) was added into 0.9 mL of NaOMe/MeOH (255mM, to form 4) or NaOEt/EtOH (312 mM, to form 5) on ice. After mixedwell, the reaction was stored at −20° C. overnight. The reaction wasquenched with addition of acetic acid to pH=7. After organic solvent wasremoved in vacuum, the residue was dissolved in water, and extractedwith cyclohexane to get rid of the non-polar impurity. The water layerwas then concentrated and injected into reverse HPLC for purification.

O-methyl β-nicotinic riboside (4): 1H NMR (400 MHz, D₂O): δ=9.74 (s,1H), 9.24 (d, 1H), 9.05 (d, 1H), 8.23 (t, 1H), 6.21 (d, 1H), 4.42 (m,2H), 4.29 (t, 1H), 3.99 (m, 4H), 3.84 (dd, 1H). 13C NMR (300 MHz, D₂O):δ=147.8, 144.0, 142.2, 129.0, 100.5, 87.9, 77.8, 69.9, 60.4, 54.4. MS:m/z (%): 270.1 (42, [M]*), 152.1 (81), 138.0 (92). HRMS (ESI) m/z [M]⁺calcd for C₁₂H₁₆NO₆ 270.09721, found 270.09737.

O-ethyl β-nicotinic riboside (5): 1H NMR (400 MHz, D₂O): δ=9.86 (s, 1H),9.37 (d, 1H), 9.20 (d, 1H), 8.37 (t, 1H), 6.35 (d, 1H), 4.55 (m, 4H),4.44 (t, 1H), 4.15 (dd, 1H), 3.98 (dd, 1H), 1.49 (t, 3H). 13C NMR (300MHz, D₂O): δ=148.9, 145.0, 143.5, 129.8, 101.7, 88.8, 79.0, 70.8, 65.7,61.4, 15.0. MS: m/z (%): 284.1 (77, [M]⁺), 256.0 (6), 212.0 (83), 133.0(7), 124.0 (14). HRMS (ESI) m/z [M]⁺ calcd for C₁₃H₁₈NO₆ 284.11286,found 284.11339.

β-Nicotinic Acid Riboside (6)

Compound 5 was dissolved into phosphate buffer (150 mM, pH=7.0). 10 uLof esterase was added to the mixture, and mixed well. The reaction wasrun at 25° C. overnight. HPLC injection showed the purity of product isabove 98%. 1H NMR (400 MHz, D₂O): δ=; MS: m/z (%): 256.1 (3, [M]⁺),228.0 (80), 207.1 (42), 146.1 (17), 124.0 (23).

N-Alkyl β-NAR Derivatives (9, 10, 11, 12, 13):

Compound 8 (25 mg, 0.61 μmol) was added to various amine (400 μM) in 0.9mL of trifluoroethanol. After mixed well, the reaction was stored at 4°C. overnight. On the next day, 4 N of NH₃/MeOH was added into thereaction mixture, and the reaction was stored at −20° C. overnight.After the reaction was quenched with addition of HCl to make pH<7, theorganic solvent was removed in vacuum. The residue was dissolved inwater and isolated with an Octadecyl-C18 disposable extraction column.

N-methyl β-nicotinamide riboside (9): 1H NMR (400 MHz, D₂O): δ=9.58 (s,1H), 9.28 (d, 1H), 8.95 (d, 1H), 8.30 (t, 1H), 6.27 (d, 1H), 4.75 (1H),4.53 (m, 1H), 4.39 (t, 1H), 4.08 (dd, 1H), 3.93 (dd, 1H), 3.00 (s, 3H).13C NMR (300 MHz, D₂O): δ=145.3, 142.4, 140.0, 128.4, 100.0, 87.8, 77.5,69.9, 60.2, 27.5. MS: m/z (%): 269.1 (13, [M]⁺), 248.0, 212.0, 137.0(100). HRMS (ESI) m/z [M]⁺ calcd for C₁₂H₁₇N₂O₅ 269.11320, found269.11378.

N-ethyl β-nicotinamide riboside (10): 1H NMR (400 MHz, D₂O): δ=9.57 (s,1H), 9.27 (d, 1H), 8.95 (d, 1H), 8.29 (t, 1H), 6.27 (d, 1H), 4.52 (m,2H), 4.38 (t, 1H), 4.07 (dd, 1H), 3.92 (dd, 1H), 3.49 (q, 2H), 1.26 (t,3H). 13C NMR (300 MHz, D₂O): δ=145.8, 142.7, 140.3, 129.0, 100.5, 88.0,77.8, 70.0, 60.6, 36.0, 13.4. MS: m/z (%): 283.1 (93, [M]⁺), 227 (50),207.1 (100), 151.1 (44).

N-allyl β-nicotinamide riboside (11): 1H NMR (400 MHz, D₂O): δ=9.59 (s,1H), 9.28 (d, 1H), 8.98 (d, 1H), 8.30 (t, 1H), 6.27 (d, 1H), 5.99 (m,1H), 5.27 (m, 1H), 4.52 (m, 2H), 4.38 (t, 1H), 4.08 (m, 3H), 3.92 (dd,1H). 13C NMR (300 MHz, D₂O): δ=146.2, 143.5, 140.7, 133.7, 129.4, 117.3,100.9, 88.4, 78.2, 70.8, 61.0, 43.5. MS: m/z (%): 295.1 (63, [M]⁺),163.1 (98), 133.1 (5), 123.1 (88).

N-ethanol β-nicotinamide riboside (12): 1H NMR (400 MHz, D₂O): δ=9.55(s, 1H), 9.24 (d, 1H), 8.93 (d, 1H), 8.24 (t, 1H), 6.22 (d, 1H), 5.99(m, 1H), 4.48 (m, 2H), 4.33 (t, 1H), 4.02 (dd, 1H), 3.87 (dd, 1H), 3.77(t, 2H), 3.58 (t, 2H). 13C NMR (300 MHz, D₂O): δ=145.0, 142.7, 140.3,128.2, 99.3, 88.0, 77.8, 69.6, 59.9, 48.5, 41.9. MS: m/z (%): 295.1 (63,[M]⁺), 163.1 (98), 133.1 (5), 123.1 (88).

N-Dimethyl β-NAR:

MS: m/z (%): 283.3 (13, [M]⁺), 151.2 (100).

5′-O-acetyl N-alkyl β-NAR derivatives (9′,10′, 11′,12′, 13′)

Compound 8 (25 mg, 0.61 μmol) was added to various amine (400 μM) in 0.9mL of trifluoroethanol. After mixed well, the reaction was stored at 4°C. overnight. After the reaction was quenched with addition of HCl tomake pH<7, the organic solvent was removed in vacuum. The residue wasdissolved in water and isolated with an Octadecyl-C18 disposableextraction column.

5′-O-acetyl N-methyl β-nicotinic riboside (9′): 1H NMR (400 MHz, D₂O)δ=9.42 (s, H_(2′), 1H), 9.23 (d, H₆, 1H), 8.98 (d, H₄, 1H), 8.33 (t, H₅,1H), 6.30 (d, H_(1′), 1H), 4.73 (t, H_(2′), 1H), 4.55 (m, 3H), 4.40 (t,H_(3′), 1H), 3.02 (s, 3H), 2.11 (s, 3H). MS: m/z (%): 311.1 (67, [M]⁺),269.1 (13), 221.1 (13), 137.1 (100).

5′-O-acetyl N-ethyl β-nicotinic riboside (10′): MS: m/z (%): 325.1 (12,[M]⁺), 283.1 (7), 151.1 (77), 139.1 (22), 115.1 (68).

5′-O-acetyl N-allyl β-nicotinic riboside (11′): MS: m/z (%): 337.1 (18,[M]⁺), 163.0 (71), 138.0 (100), 124.0 (18).

5′-O-ethylenediamine N-methyl β-nicotinic riboside (12′): MS: m/z (%):340.2 (100, [M]⁺), 298.1 (44), 166.1 (100), 124.0 (82).

5′-O-acetyl N-dimethyl β-nicotinamide riboside (300 MHz, D₂O) δ=9.23 (s,H₂, 1H), 9.19 (d, H₆, 1H), 8.78 (d, H₄, 1H), 8.32 (t, H₅, 1H), 6.28 (d,H_(1′), 1H), 4.72 (m, H_(4′), 1H), 4.55 (t, H_(2′), 1H), 4.52 (d,H_(5′), 2H), 4.40 (t, H_(3′), 1H), 3.17 (s, 3H), 3.08 (s, 3H), 2.10 (s,3H). MS: m/z (%): 325.1 (92, [M]⁺), 283.1 (2) 151.1 (100).

1. A method for increasing intracellular levels of nicotinamide adeninedinucleotide (NAD+) in cells, the method comprising administering to thecells a therapeutically effective amount of a compound of formula (I)

and salt complexes thereof, wherein R is selected from the groupconsisting of alkylamino, substituted alkylamino, dialkylamino,substituted dialkylamino, alkyloxy, substituted alkyloxy, aryloxy,substituted aryloxy, hydroxylamino, substituted hydroxylamino,O-alkyloxyamino, substituted O-alkyloxyamino, aminooxy, substitutedaminooxy, N-alkylaminooxy, substituted N-alkylaminooxy, hydrazino,substituted hydrazino, alkylhydrazino, and substituted alkylhydrazino,wherein the administering increases the intracellular levels of NAD+ inthe cells.
 2. The method of claim 1, wherein R is alkylamino orsubstituted alkylamino.
 3. The method of claim 2, wherein the alkylaminoor substituted alkylamino is —NHMe, —NHEt, —NHCH₂CH₂OH, —NHCH₂CH═CH₂,—NHCH(CH₃)₂, —NHCH₂CH₂NH₂, or —NHcyclopropyl.
 4. The method of claim 1,wherein R is dialkylamino or substituted dialkylamino.
 5. The method ofclaim 4, wherein the dialkylamino or substituted dialkylamino is—N(CH₃)₂ or N-substituted pyrrolidinyl.
 6. The method of claim 1,wherein R is alkyloxy or substituted alkyloxy.
 7. The method of claim 6,wherein the alkyloxy or substituted alkyloxy is —OMe, —OEt, or—OCH₂CH₂OH.
 8. The method of claim 1, wherein R is aryloxy orsubstituted aryloxy.
 9. The method of claim 8, wherein the aryloxy orsubstituted aryloxy is —OPh.
 10. The method of claim 1, wherein R ishydroxylamino, substituted hydroxylamino, O-alkyloxyamino, substitutedO-alkyloxyamino, aminooxy, substituted aminooxy, N-alkylaminooxy,substituted N-alkylaminooxy, hydrazino, substituted hydrazino,alkylhydrazino, or substituted alkylhydrazino.
 11. The method of claim1, wherein the compound of formula (I) is in the form of apharmaceutical composition or a nutritional supplement.
 12. The methodof claim 1, wherein the administering increases the intracellular levelsof NAD+ in vivo.
 13. The method of claim 1, wherein the administeringincreases the intracellular levels of NAD+ex vivo.
 14. The method ofclaim 1, wherein the administering comprises topical administration. 15.The method of claim 1, wherein the administering comprises intravenousadministration.
 16. The method of claim 1, wherein the administeringcomprises oral administration.
 17. The method of claim 1, wherein theadministering comprises inhalation or insufflation administration. 18.The method of claim 1, wherein the administering is performed on arecipient who has received, or is to receive, a dose of radiation ortoxin.
 19. The method of claim 1, wherein the administering comprises acombination therapeutic comprising a cardiovascular agent.
 20. Themethod of claim 19, wherein the cardiovascular agent comprises ananti-arrhythmic agent, an antihypertensive agent, a calcium channelblocker, a cardioplegic solution, a cardiotonic agent, a fibrinolyticagent, a sclerosing solution, a vasoconstrictor agent, a vasodilatoragent, a nitric oxide donor, a potassium channel blocker, a sodiumchannel blocker, statins, or a naturiuretic agent.